Method and process for the production of multi-coated recognitive and releasing systems

ABSTRACT

The present invention includes compositions, methods, systems for the controlled delivery of an active agent within a polymeric network upon the binding of a molecule that decreases the structural integrity of the polymeric network at one or more micro- or nanovacuoles.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. Provisional Application Ser.No. 60/894,451, filed Mar. 12, 2007, and is a continuation-in-part ofU.S. patent application Ser. No. 12/047,309, filed Mar. 12, 2008, theentire contents of which are incorporated herein by reference.

TECHNICAL FIELD OF THE INVENTION

The present invention relates in general to the field of the controlledrelease of agents, and more particularly, to novel compositions andmethods for making controlled release configurational biomimeticimprinting networks.

STATEMENT OF FEDERALLY FUNDED RESEARCH

None.

BACKGROUND OF THE INVENTION

Without limiting the scope of the invention, its background is describedin connection with the recognition and controlled release of activeagents from polymers.

U.S. Pat. No. 7,459,316, issued to Faid, et al., is directed to aMolecularly-Imprinted Chemical Detection Device and Method. Briefly, anovel method of molecular imprinting is described that uses a modifiedsoft lithography technique, a molecularly-imprinted chemical detectiondevice comprising at least one molecularly-imprinted polymer capable ofdetecting at least one chemical target is produced. The device can beused in the field for in situ detection and quantification of chemicaltargets using standard surface analytical techniques.

U.S. Pat. No. 7,176,247, issued to Walker, teaches an interpenetratingpolymer network. Briefly, a water insoluble interpenetrating polymernetwork is obtained by independently cross-linking a first polymerderived from a sulfonic acid or phosphonic acid group containing alkenylmonomer and a second polymer polymerized independently of the firstpolymer and interpenetrating the first polymer, where the second polymeris selectively permeable to water compared to methanol. Throughadjustment of the degree of first polymer monomer acidification, polymerratios and the extent of cross-linking in the at least twointerpenetrating polymers, ion conductivity and solvent permeability arecontrolled. The relative degree and mechanism of cross-linking andinterpenetrating the first polymer and second polymer are alsoadjustable parameters that impact on film properties.

United States Patent Application No. 20080171067, filed by Serengulam,et al., is directed to Polymeric Carriers of Therapeutic Agents andRecognition Moieties for Antibody-Based Targeting of Disease Sites.Briefly, the disclosure teaches methods and compositions for delivery oftherapeutic agents to target cells, tissues or organisms. In preferredembodiments, the therapeutic agents are delivered in the form oftherapeutic-loaded polymers that may comprise many copies of one or moretherapeutic agents. The polymer may be conjugated to a peptide moietythat contains one or more haptens, such as HSG. Theagent-polymer-peptide complex may be delivered to target cells by, forexample, a pre-targeting technique utilizing bispecific or multispecificantibodies or fragments, having at least one binding arm that recognizesthe hapten and at least a second binding arm that binds specifically toa disease or pathogen associated antigen, such as a tumor associatedantigen. Methods for synthesizing and using such therapeutic-loadedpolymers and their conjugates are provided.

SUMMARY OF THE INVENTION

The needs of the invention set forth above as well as further and otherneeds and advantages of the present invention are achieved by theembodiments of the invention described herein below. The presentinvention is based on the recognition that, to date, imprinted orrecognitive polymers are found in two forms, solid and gel-like. Solidrecognitive polymers are used in a variety of applications, namely,chromatography, filters and molecular separation. The other category ofimprinted polymers are gelatinous polymers that are loaded with apayload during the polymerization phase and are dried until use. Uponexposing these gelatinous imprinted polymers to a solvent, e.g., waterand/or water with the analyte or recognitive molecules, the gelatinouspolymers swell. Unfortunately, for the delivery of most payloadsswelling of the gelatinous polymer in the presence of solvent aloneleads to leakage of the payload. What are needed are compositions,methods and systems for the delivery of payloads under recognitivecontrol that do not leak and that are amenable to controlled releaseupon exposure to the analyte.

The present invention also includes a method of making a polymericrecognitive network that includes selecting one or more targets forrecognition; forming micro- or nano-vacuoles in the polymericrecognitive network about the one or more targets; embedding within thepolymeric recognitive network one or more active agents for release upondissociation of the polymeric recognitive network; and removing thetargets from the micro- or nano-vacuoles, wherein subsequent binding ofthe target to the micro- or nano-vacuoles causes disruption of thepolymeric recognitive network and release of the one or more activeagents.

Yet another embodiment of the present invention includes a kit formaking a polymeric recognitive network that includes one or more targetsfor recognition by the polymeric recognitive network; monomers forforming a polymeric recognitive network comprising micro- ornano-vacuoles about the one or more targets; a polymeric catalyst forforming the polymeric recognitive network about the targets; andinstructions for polymerizing the polymeric recognitive network andremoving the targets from the micro- or nano-vacuoles and loading of thepolymeric recognitive network with one or more active agents.

According to one embodiment, the present disclosure provides a systemfor forming multilayer mimetic structures that comprise a core andcoating materials, including molecularly imprinted polymers, materialsfor use as a spacer, and materials for use as a binder. According toanother embodiment, the present disclosure provides a system for formingmultilayer mimetic structures wherein the core comprises spherical ornon-spherical compositions, molecularly imprinted polymers, hydroxylpropyl cellulose (HPC) as a spacer and mannitol as a binder. Accordingto another embodiment, the present disclosure provides a system forforming multilayer mimetic structures in which the molecularly imprintedpolymer layer with controllable properties for bursting and drugrelease.

According to another embodiment, the present disclosure provides methodsfor controlling bursting and release of molecularly imprinted polymerlayers by manipulation of various factors including, but not limited to,disintegration kinetics, tensile strength of the polymers viacrosslinking and intrapolymer complexes, molecular weight, pressuredifferences by use of osmotic agents, and thickness of the layer.

The present invention includes a molecule-imprinted polymeric networkthat includes a polymer or gel comprising one or more micro- ornanovacuoles, or micro- or nanopores wherein the nanovacuoles ornanopores recognize a specific molecule while subsequent contact withthe molecule creates internal stresses that rupture the polymericnetwork at the micro-or nanovacuoles or micro- or nanopores. Uponexposure of the polymeric network to the analyte, but not the solventalone, the polymeric network can rupture due to, e.g., osmosis uponrecognition and binding of the molecule leading to rupture due toswelling; change of the solubility of the polymeric network leading topolymer dissolution; local temperature changes leading to expansion ofthe polymeric network and combinations thereof. The composition may beloaded with one or more active agents to form an active agent-loaded,molecule-imprinted network. In one aspect of the invention, the polymerswells between 2-20, 4-18, 5-15, 8-12 and 10 percent of the driedpolymer upon exposure to the solvent alone. In one aspect, the polymericnetwork swell between 5-15% of the dried polymeric network in thepresence of the solvent alone.

Examples of active agents for use with the present invention includepharmaceutical and medical applications, food components, detergents,bleaches, fabric softeners, fragrances, cosmetic products, airfresheners, room deodorant devices, perfumed substrates, perfumedplastics and pet collars. Other actives include food and cosmeticapplications that use hydrocolloids as imprinting carriers for polymersof high molecular weight, wherein the hydrocolloids are extracted fromplants, seaweeds or animal collagen, produced by microbial synthesis,and comprise polysaccharides, proteins and combinations thereof. Themolecule-imprinted network may be a carbohydrate polymer of glycosidictype mono-sugar repeative units, galactomannans, pectins, alginates,carrageenans and xanthan gum that are linear or branched, neutral oranionic and combinations thereof. Other examples include householdproducts selected from laundry care; paper products; specialty cleaners(chlorinated cleaners, scouring pads, effervescent toilet bowl cleanerpowders); air fresheners and combinations thereof. Further examplesinclude personal care products selected from hair care (shampoos, hairmousses, styling agents); skin care (body lotions, vitamin e, aloevera); bath products (moisture-triggered release products); bodypowders; toilet soap (milled or poured, ionic strength-triggeredrelease) and combinations thereof. Also, cosmetics and treatmentproducts selected from lipstick; eye-liner; foundation; base; blush;mascara; eye shadow; lip liner; facial powder; consealer; facial cream;make-up remover; mascara remover; make-up; skin treatments and may eveninclude one or more fragrances or carriers therefore that includecologne; perfume; sampling; antiperspirant; deodorant; anti-dandruffshampoos; athlete foot products and combinations thereof. Other activesinclude a surfactant, a bleaching agent, a corrosion inhibitor, asudsing modifier, a fluorescent whitening agent, one or more enzymes, ananti redeposition agent, a color, a fragrance, one or more additives andcombinations thereof.

The present invention also include the release of active agent uponexposure to the cognate molecule or moiety that ultimately causesrelease upon a loss of structural integrity caused by, e.g., changes insolubility, pressure, a pH shift, a change in temperature, a temperatureincrease, enzymatic breakdown, diffusion and combinations thereof. Theskilled artisan will also be able to be formed integrally or as acoating for controlled release or one or more layers of the active, therecognitive polymer or both. Furthermore, the polymeric recognitivenetwork may be formed into one or more layers, each of which recognizesa different molecule, a different active or inert agent or both. Thepolymeric recognitive network is formed into a sphere, film, planar,semi-spherical, cylinder, rod, hemispheres, conical, hemi-cylinders andcombination thereof and may also be at least partially porous. Thepolymeric recognitive network also may be formed into one or morelayers, each of which recognizes a different molecule, and where adifferent active or inert agent may be contained between polymericlayers.

The present invention also includes an active agent-loaded,molecule-imprinted polymeric network that includes two or more activeagent loaded, aggregated polymeric or gel nanoparticles ormicroparticles comprising micro- or nanovacuoles or micro- or nanoporespreviously imprinted with a molecule, wherein one or more pre-determinedmolecules bind specifically to the micro- or nanovacuoles or micro- ornanopores and contact with the molecule creates internal stresses thatrupture the network at the micro- or nanovacuoles or micro- or nanoporesthereby releasing one or more active agents loaded into the activeagent-loaded, molecule-imprinted polymeric network.

The present invention also includes a method of making a recognitiverelease system by selecting one or more molecules for recognition;forming micro- or nano-vacuoles in a polymeric recognitive network aboutthe one or more molecules; removing the molecule from the micro- ornanovacuoles or micro- or nanopores; and coating one or more activeagents with a polymeric recognitive network, wherein the one or moreactive agents release upon contact by the polymeric recognitive networkwith its cognate molecule.

BRIEF DESCRIPTION OF THE DRAWINGS

For a more complete understanding of the features and advantages of thepresent invention, reference is now made to the detailed description ofthe invention along with the

FIG. 1 shows the monomeric mixture used to create the present invention.

FIG. 2 shows the polymerized mixture, before the washing step.

FIG. 3 shows the polymer after washing in which the template has beenremoved and the specific recognition site remains within the polymer.

FIG. 4 is a graph that shows penetrant uptake of a recognitive polymercontinuous films over time. The data points are amount of penetrantuptake in Milli-Q deionized water (DI water) and 100 mg/dL D-glucose andDI water. The films were cut into disks 8 mm in diameter and 0.12 mmthick; the initial weights were approximately 6 mg each. Measurementswere taken every 10 minutes.

FIG. 5 is a graph that shows penetrant uptake of a recognitive polymercontinuous film versus the square root of time. The data points areamount of penetrant uptake in Milli-Q deionized water (DI water) and 100mg/dL D-glucose and DI water. The films were cut into disks 8 mm indiameter and 0.12 mm thick; the weights were approximately 6 mg each.Measurements were taken every 10 minutes.

FIG. 6 is a graph that shows the recognitive ratio of configurationalbiomimetic imprinted polymers (CBIP) in the presence of 100 mg per dLdeionized water compared to continuous films in the presence ofdeionized water. The films were cut into squares approximately 9 mm by 9mm and 0.22 mm thick. The penetrant uptake amount was obtained frommeasurements of mass every 10 minutes once the squares were placed inthe solutions of either deionized Water or glucose solution with 100 mgD-glucose per dL deionized water. The ratio is the amount of penetrantuptake in the glucose solution to amount of penetrant uptake indeionized water.

FIG. 7 shows a Glucose CBIP (5/31 mixture) after exposure to 100 mg/dLglucose-water (60 seconds pass between each frame). (50× objective).[0026] FIG. 8 shows a nebula of disintegrated particles from a porousfilm exposed to 100 mg/dL glucose-water (5× objective).

FIG. 9 shows the Stress lines seen on a polymer film 30 sec afteraddition of glucose. (5× objective).

FIG. 10 shows a section of a glucose-CBIP film immersed in a glucosesolution and seen breaking at a first time point (10× objective).

FIG. 11 shows a section of a glucose-CBIP film immersed in a glucosesolution and seen breaking at a second time point (10× objective).

FIG. 12 shows a Trypan Blue dyed water-front moving in toward a CBIPparticle.

FIG. 13 shows a CBIP particle in glucose-water viewed through twopolarized lenses. Soft image means no significant stresses in thesurface of the particle (the image is mostly dark due to the lack ofreflection).

FIG. 14 shows a CBIP polymer particle viewed through two polarizedlenses. The reflective areas indicate changes in surface morphology dueto mechanical stresses.

FIG. 15 shows a glucose CBIP exposed to fluorescent glucose; the intakeof the glucose is indicated by the brightly lit areas (10× objective).

FIG. 16 shows a glucose CBIP exposed to fluorescent glucose; breakage isapparent here (10× objective).

FIG. 17 shows the optional combinations for the various embodiments ofthe present invention in which the optional analyte, recognition andtransduction events and payloads.

FIG. 18 is a diagram that shows mixing multilayered mimetic structureswith different release profiles that allows the system to be tailored tofit any release profiles. Mixing four different microcapsules allowedthe system to rupture every 3 h for 4 days instead of a system thatruptured every 12 h for 4 days or a system that ruptured every 3 h forone day.

FIG. 19 shows glucose/water uptake of a CBIP.

DETAILED DESCRIPTION OF THE INVENTION

While the making and using of various embodiments of the presentinvention are discussed in detail below, it should be appreciated thatthe present invention provides many applicable inventive concepts thatcan be embodied in a wide variety of specific contexts. The specificembodiments discussed herein are merely illustrative of specific ways tomake and use the invention and do not delimit the scope of theinvention.

To facilitate the understanding of this invention, a number of terms aredefined below. Terms defined herein have meanings as commonly understoodby a person of ordinary skill in the areas relevant to the presentinvention. Terms such as “a”, “an” and “the” are not intended to referto only a singular entity, but include the general class of which aspecific example may be used for illustration. The terminology herein isused to describe specific embodiments of the invention, but their usagedoes not delimit the invention, except as outlined in the claims.

As used herein, the term “active agent(s),” “active ingredient(s),”“pharmaceutical ingredient(s),” and “bioactive agent(s)” are defined asdrugs and/or pharmaceutically active ingredients. The present inventionmay be used to encapsulate, attach, bind or otherwise be used to affectthe storage, stability, longevity and/or release of any of the followingdrugs as the pharmaceutically active agent in a composition. One or moreof the following bioactive agents may be combined with one or morecarriers and the present invention (which may itself be the carrier):

Analgesic anti-inflammatory agents such as, acetaminophen, aspirin,salicylic acid, methyl salicylate, choline salicylate, glycolsalicylate, 1-menthol, camphor, mefenamic acid, fluphenamic acid,indomethacin, diclofenac, alclofenac, ibuprofen, ketoprofen, naproxene,pranoprofen, fenoprofen, sulindac, fenbufen, clidanac, flurbiprofen,indoprofen, protizidic acid, fentiazac, tolmetin, tiaprofenic acid,bendazac, bufexamac, piroxicam, phenylbutazone, oxyphenbutazone,clofezone, pentazocine, mepirizole, and the like.

Drugs having an action on the central nervous system, for examplesedatives, hypnotics, antianxiety agents, analgesics and anesthetics,such as, chloral, buprenorphine, naloxone, haloperidol, fluphenazine,pentobarbital, phenobarbital, secobarbital, amobarbital, cydobarbital,codeine, lidocaine, tetracaine, dyclonine, dibucaine, cocaine, procaine,mepivacaine, bupivacaine, etidocaine, prilocaine, benzocaine, fentanyl,nicotine, and the like. Local anesthetics such as, benzocaine, procaine,dibucaine, lidocaine, and the like.

Antihistaminics or antiallergic agents such as, diphenhydramine,dimenhydrinate, perphenazine, triprolidine, pyrilamine, chlorcyclizine,promethazine, carbinoxamine, tripelennamine, brompheniramine,hydroxyzine, cyclizine, meclizine, clorprenaline, terfenadine,chlorpheniramine, and the like. Anti-allergenics such as, antazoline,methapyrilene, chlorpheniramine, pyrilamine, pheniramine, and the like.Decongestants such as, phenylephrine, ephedrine, naphazoline,tetrahydrozoline, and the like.

Antipyretics such as, aspirin, salicylamide, non-steroidalanti-inflammatory agents, and the like. Antimigrane agents such as,dihydroergotamine, pizotyline, and the like. Acetonide anti-inflammatoryagents, such as hydrocortisone, cortisone, dexamethasone, fluocinolone,triamcinolone, medrysone, prednisolone, flurandrenolide, prednisone,halcinonide, methylprednisolone, fludrocortisone, corticosterone,paramethasone, betamethasone, ibuprophen, naproxen, fenoprofen,fenbufen, flurbiprofen, indoprofen, ketoprofen, suprofen, indomethacin,piroxicam, aspirin, salicylic acid, diflunisal, methyl salicylate,phenylbutazone, sulindac, mefenamic acid, meclofenamate sodium,tolmetin, and the like. Muscle relaxants such as, tolperisone, baclofen,dantrolene sodium, cyclobenzaprine.

Steroids such as, androgenic steriods, such as, testosterone,methyltestosterone, fluoxymesterone, estrogens such as, conjugatedestrogens, esterified estrogens, estropipate, 17-β estradiol, 17-βestradiol valerate, equilin, mestranol, estrone, estriol, 17β ethinylestradiol, diethylstilbestrol, progestational agents, such as,progesterone, 19-norprogesterone, norethindrone, norethindrone acetate,melengestrol, chlormadinone, ethisterone, medroxyprogesterone acetate,hydroxyprogesterone caproate, ethynodiol diacetate, norethynodrel, 17-αhydroxyprogesterone, dydrogesterone, dimethisterone, ethinylestrenol,norgestrel, demegestone, promegestone, megestrol acetate, and the like.

Respiratory agents such as, theophilline and β2-adrenergic agonists,such as, albuterol, terbutaline, metaproterenol, ritodrine, carbuterol,fenoterol, quinterenol, rimiterol, solmefamol, soterenol, tetroquinol,and the like. Sympathomimetics such as, dopamine, norepinephrine,phenylpropanolamine, phenylephrine, pseudoephedrine, amphetamine,propylhexedrine, arecoline, and the like.

Antimicrobial agents including antibacterial agents, antifungal agents,antimycotic agents and antiviral agents; tetracyclines such as,oxytetracycline, penicillins, such as, ampicillin, cephalosporins suchas, cefalotin, aminoglycosides, such as, kanamycin, macrolides such as,erythromycin, chloramphenicol, iodides, nitrofrantoin, nystatin,amphotericin, fradiomycin, sulfonamides, purrolnitrin, clotrimazole,miconazole chloramphenicol, sulfacetamide, sulfamethazine, sulfadiazine,sulfamerazine, sulfamethizole and sulfisoxazole; antivirals, includingidoxuridine; clarithromycin; and other anti-infectives includingnitrofurazone, and the like.

Antihypertensive agents such as, clonidine, α-methyldopa, reserpine,syrosingopine, rescinnamine, cinnarizine, hydrazine, prazosin, and thelike. Antihypertensive diuretics such as, chlorothiazide,hydrochlorothrazide, bendoflumethazide, trichlormethiazide, furosemide,tripamide, methylclothiazide, penfluzide, hydrothiazide, spironolactone,metolazone, and the like. Cardiotonics such as, digitalis,ubidecarenone, dopamine, and the like. Coronary vasodilators such as,organic nitrates such as, nitroglycerine, isosorbitol dinitrate,erythritol tetranitrate, and pentaerythritol tetranitrate, dipyridamole,dilazep, trapidil, trimetazidine, and the like. Vasoconstrictors suchas, dihydroergotamine, dihydroergotoxine, and the like. β-blockers orantiarrhythmic agents such as, timolol pindolol, propranolol, and thelike. Humoral agents such as, the prostaglandins, natural and synthetic,for example PGE1, PGE2α, and PGF2α, and the PGE1 analog misoprostol.Antispasmodics such as, atropine, methantheline, papaverine,cinnamedrine, methscopolamine, and the like.

Calcium antagonists and other circulatory organ agents, such as,aptopril, diltiazem, nifedipine, nicardipine, verapamil, bencyclane,ifenprodil tartarate, molsidomine, clonidine, prazosin, and the like.Anti-convulsants such as, nitrazepam, meprobamate, phenytoin, and thelike. Agents for dizziness such as, isoprenaline, betahistine,scopolamine, and the like. Tranquilizers such as, reserprine,chlorpromazine, and antianxiety benzodiazepines such as, alprazolam,chlordiazepoxide, clorazeptate, halazepam, oxazepam, prazepam,clonazepam, flurazepam, triazolam, lorazepam, diazepam, and the like.

Antipsychotics such as, phenothiazines including thiopropazate,chlorpromazine, triflupromazine, mesoridazine, piperracetazine,thioridazine, acetophenazine, fluphenazine, perphenazine,trifluoperazine, and other major tranqulizers such as, chlorprathixene,thiothixene, haloperidol, bromperidol, loxapine, and molindone, as wellas, those agents used at lower doses in the treatment of nausea,vomiting, and the like.

Drugs for Parkinson's disease, spasticity, and acute muscle spasms suchas levodopa, carbidopa, amantadine, apomorphine, bromocriptine,selegiline (deprenyl), trihexyphenidyl hydrochloride, benztropinemesylate, procyclidine hydrochloride, baclofen, diazepam, dantrolene,and the like. Respiratory agents such as, codeine, ephedrine,isoproterenol, dextromethorphan, orciprenaline, ipratropium bromide,cromglycic acid, and the like. Non-steroidal hormones or antihormonessuch as, corticotropin, oxytocin, vasopressin, salivary hormone, thyroidhormone, adrenal hormone, kallikrein, insulin, oxendolone, and the like.

Vitamins such as, vitamins A, B, C, D, E and K and derivatives thereof,calciferols, mecobalamin, and the like for dermatologically use. Enzymessuch as, lysozyme, urokinaze, and the like. Herb medicines or crudeextracts such as, Aloe vera, and the like.

Antitumor agents such as, 5-fluorouracil and derivatives thereof,krestin, picibanil, ancitabine, cytarabine, and the like. Anti-estrogenor anti-hormone agents such as, tamoxifen or human chorionicgonadotropin, and the like. Miotics such as pilocarpine, and the like.

Cholinergic agonists such as, choline, acetylcholine, methacholine,carbachol, bethanechol, pilocarpine, muscarine, arecoline, and the like.Antimuscarinic or muscarinic cholinergic blocking agents such as,atropine, scopolamine, homatropine, methscopolamine, homatropinemethylbromide, methantheline, cyclopentolate, tropicamide,propantheline, anisotropine, dicyclomine, eucatropine, and the like.

Mydriatics such as, atropine, cyclopentolate, homatropine, scopolamine,tropicamide, eucatropine, hydroxyamphetamine, and the like. Psychicenergizers such as 3-(2-aminopropy)indole, 3-(2-aminobutyl)indole, andthe like.

Antidepressant drugs such as, isocarboxazid, phenelzine,tranylcypromine, imipramine, amitriptyline, trimipramine, doxepin,desipramine, nortriptyline, protriptyline, amoxapine, maprotiline,trazodone, and the like.

Anti-diabetics such as, insulin, and anticancer drugs such as,tamoxifen, methotrexate, and the like.

Anorectic drugs such as, dextroamphetamine, methamphetamine,phenylpropanolamine, fenfluramine, diethylpropion, mazindol,phentermine, and the like.

Anti-malarials such as, the 4-aminoquinolines, alphaaminoquinolines,chloroquine, pyrimethamine, and the like.

Anti-ulcerative agents such as, misoprostol, omeprazole, enprostil, andthe like. Antiulcer agents such as, allantoin, aldioxa, alcloxa,N-methylscopolamine methylsuflate, and the like. Antidiabetics such asinsulin, and the like.

Anti-cancer agent such as, cis-platin, actinomycin D, doxorubicin,vincristine, vinblastine, etoposide, amsacrine, mitoxantrone,tenipaside, taxol, colchicine, cyclosporin A, phenothiazines orthioxantheres.

For use with vaccines, one or more antigens, such as, natural,heat-killer, inactivated, synthetic, peptides and even T cell epitopes(e.g., GADE, DAGE, MAGE, etc.) and the like.

Example therapeutic or active agents also include water soluble orpoorly soluble drug of molecular weigh from 40 to 1,100 including thefollowing: Hydrocodone, Lexapro, Vicodin, Effexor, Paxil, Wellbutrin,Bextra, Neurontin, Lipitor, Percocet, Oxycodone, Valium, Naproxen,Tramadol, Ambien, Oxycontin, Celebrex, Prednisone, Celexa, Ultracet,Protonix, Soma, Atenolol, Lisinopril, Lortab, Darvocet, Cipro, Levaquin,Ativan, Nexium, Cyclobenzaprine, Ultram, Alprazolam, Trazodone, Norvasc,Biaxin, Codeine, Clonazepam, Toprol, Zithromax, Diovan, Skelaxin,Klonopin, Lorazepam, Depakote, Diazepam, Albuterol, Topamax, Seroquel,Amoxicillin, Ritalin, Methadone, Augmentin, Zetia, Cephalexin, Prevacid,Flexeril, Synthroid, Promethazine, Phentermine, Metformin, Doxycycline,Aspirin, Remeron, Metoprolol, Amitriptyline, Advair, Ibuprofen,Hydrochlorothiazide, Crestor, Acetaminophen, Concerta, Clonidine, Norco,Elavil, Abilify, Risperdal, Mobic, Ranitidine, Lasix, Fluoxetine,Coumadin, Diclofenac, Hydroxyzine, Phenergan, Lamictal, Verapamil,Guaifenesin, Aciphex, Furosemide, Entex, Metronidazole, Carisoprodol,Propoxyphene, Digoxin, Zanaflex, Clindamycin, Trileptal, Buspar, Keflex,Bactrim, Dilantin, Flomax, Benicar, Baclofen, Endocet, Avelox, Lotrel,Inderal, Provigil, Zantac, Fentanyl, Premarin, Penicillin, Claritin,Reglan, Enalapril, Tricor, Methotrexate, Pravachol, Amiodarone, Zelnorm,Erythromycin, Tegretol, Omeprazole, and Meclizine.

The drugs mentioned above may be used in combination as required.Moreover, the above drugs may be used either in the free form or, ifcapable of forming salts, in the form of a salt with a suitable acid orbase. If the drugs have a carboxyl group, their esters may be employed.

Examples of monomers that may be used to achieve the low or minimalswelling include: Poly(allylamine), Acrylic acid, Acrylamide,(Diethylamino)ethyl methacrylate, (Ethylamino)methacrylate, Methacrylicacid, methylmethacrylate, Triazacyclononane-copper(II) complex,2-(methacryloyxloxy) ethyl phosphate, methacrylamide,2-(trifluoromethyl)acrylic acid, 3-aminophenylboronic acid,poly(allylamine), o-phthalic dialdehyde, oleyl phenyl hydrogenphosphate, 4-vinylpyridine, vinylimidazole,2-acryloilamido-2,2′-methopropane sulfonic acid, Silica, organicsilanes, N-(4-vinyl)-benzyl iminodiacetic acid, Ni(II)-nitrilotriaceticacid, N-acryloyl-alanine. These monomers may be combined with one ormore crosslinkers to achieve the desired low or minimal swelling uponexposure to solvent alone that include: ethylene glycol dimethacrylate,pentaerythritol triacrylate, pentaerythritol tetraacrylate,trimethylolpropane trimethacrylate, vinyl triethoxysilane, vinyltrimethoxysilane, toluene 2,4-diisocyanate, epichlorohydrin,triglycerolate diacrylate, polystyrene surface, Propylene glycoldimethacrylate, poly(ethylene glycol) n dimethacrylate, methacrylatederived silica, acrylonitrile, N,N′-dimethylacrylamide, poly(ethyleneglycol) diacrylate. Examples of solvents that may be used to achieve lowor minimal swelling include Acetonitrile, Acetic acid, ethanol, aqueousbuffer, toluene, water, chloroform, hexane, methanol, tetrahydrofuran.

The acid mentioned above may be an organic acid, for example,methanesulfonic acid, lactic acid, tartaric acid, fumaric acid, maleicacid, acetic acid, or an inorganic acid, for example, hydrochloric acid,hydrobromic acid, phosphoric acid or sulfuric acid. The base may be anorganic base, for example, ammonia, triethylamine, or an inorganic base,for example, sodium hydroxide or potassium hydroxide. The estersmentioned above may be alkyl esters, aryl esters, aralkyl esters, andthe like. Also with sugar to release as a bitterness masking agent(sugar as the agent).

The bioactive may also be administered, e.g., parenterally,intraperitoneally, intraspinally, intravenously, intramuscularly,intravaginally, subcutaneously, or intracerebrally. Dispersions may beprepared in glycerol, liquid polyethylene glycols, and mixtures thereofand in oils. Under ordinary conditions of storage and use, thesepreparations may contain a preservative to prevent the growth ofmicroorganisms.

Pharmaceutical compositions suitable for injectable use include sterileaqueous solutions (where water soluble) or dispersions and sterilepowders for the extemporaneous preparation of sterile injectablesolutions or dispersion. In all cases, the composition must be sterileand must be fluid to the extent that easy syringability exists. It mustbe stable under the conditions of manufacture and storage and must bepreserved against the contaminating action of microorganisms such asbacteria and fungi. The carrier may be a solvent or dispersion mediumcontaining, for example, water, ethanol, poly-ol (for example, glycerol,propylene glycol, and liquid polyethylene glycol, and the like),suitable mixtures thereof, and vegetable oils.

The proper fluidity may be maintained, for example, by the use of acoating such as lecithin, by the maintenance of the required particlesize in the case of dispersion and by the use of surfactants. Preventionof the action of microorganisms may be achieved by various antibacterialand antifungal agents, for example, parabens, chlorobutanol, phenol,ascorbic acid, thimerosal, and the like. In many cases, it will bepreferable to include isotonic agents, for example, sugars, sodiumchloride, or polyalcohols such as mannitol and sorbitol, in thecomposition. Prolonged absorption of the injectable compositions may bebrought about by including in the composition an agent that delaysabsorption, for example, aluminum monostearate or gelatin.

Sterile injectable solutions may be prepared by incorporating thetherapeutic compound in the required amount in an appropriate solventwith one or a combination of ingredients enumerated above, as required,followed by filtered sterilization. Generally, dispersions are preparedby incorporating the therapeutic compound into a sterile carrier thatcontains a basic dispersion medium and the required other ingredientsfrom those enumerated above. In the case of sterile powders for thepreparation of sterile injectable solutions, the methods of preparationmay include vacuum drying, spray drying, spray freezing andfreeze-drying that yields a powder of the active ingredient (i.e., thetherapeutic compound) plus any additional desired ingredient from apreviously sterile-filtered solution thereof.

The bioactive may be orally administered, for example, with an inertdiluent or an assimilable edible carrier. The therapeutic compound andother ingredients may also be enclosed in a hard or soft shell gelatincapsule, compressed into tablets, or incorporated directly into thesubject's diet. For oral therapeutic administration, the therapeuticcompound may be incorporated with excipients and used in the form ofingestible tablets, buccal tablets, troches, capsules, elixirs,suspensions, syrups, wafers, and the like. The percentage of thetherapeutic compound in the compositions and preparations may, ofcourse, be varied as will be known to the skilled artisan. The amount ofthe therapeutic compound in such therapeutically useful compositions issuch that a suitable dosage will be obtained.

It is especially advantageous to formulate parenteral compositions indosage unit form for ease of administration and uniformity of dosage.Dosage unit form as used herein refers to physically discrete unitssuited as unitary dosages for the subjects to be treated; each unitcontaining a predetermined quantity of therapeutic compound calculatedto produce the desired therapeutic effect in association with therequired pharmaceutical carrier. The specification for the dosage unitforms of the invention are dictated by and directly dependent on (a) theunique characteristics of the therapeutic compound and the particulartherapeutic effect to be achieved, and (b) the limitations inherent inthe art of compounding such a therapeutic compound for the treatment ofa selected condition in a subject.

Aqueous compositions of the present invention comprise an effectiveamount of the nanoparticle, nanofibril or nanoshell or chemicalcomposition of the present invention dissolved and/or dispersed in apharmaceutically acceptable carrier and/or aqueous medium. Thebiological material should be extensively dialyzed to remove undesiredsmall molecular weight molecules and/or lyophilized for more readyformulation into a desired vehicle, where appropriate. The activecompounds may generally be formulated for parenteral administration,e.g., formulated for injection via the intravenous, intramuscular,subcutaneous, intralesional, and/or even intraperitoneal routes. Thepreparation of an aqueous composition that contain an effective amountof the nanoshell composition as an active component and/or ingredientwill be known to those of skill in the art in light of the presentdisclosure. Typically, such compositions may be prepared as injectables,either as liquid solutions and/or suspensions; solid forms suitable forusing to prepare solutions and/or suspensions upon the addition of aliquid prior to injection may also be prepared; and/or the preparationsmay also be emulsified.

The present inventor has developed methods and techniques to formsynthetic biomimetic networks, gels or polymers that will bind andrespond to specific molecules, analytes or moieties. These biomimeticpolymer networks, gels or polymers are advantageous because they can betailored to bind any molecule with controlled selectivity and affinity.

There are some significant characteristics to consider in the design ofa biomimetic polymer networks via a configurational biomimeticimprinting (CBIP) technique. To achieve a relatively easy on/off bindingevent, a non-covalent recognition process is favored. Therefore,supramolecular interactions, such as hydrogen bonding, electrostaticinteractions, hydrophobic interactions, and van der Waals forces, areemployed to achieve recognition. For the formation of the network, it isimperative that the functional monomers, crosslinker, and template aremutually soluble. In addition, the solvent must be chosen wisely, sothat it does not interact and destabilize the self-assembled functionalmonomer and template.

The ability to engineer traditional polymers with specific materialproperties is hampered by lack of control of molecular weight, chainconfiguration and polymerization kinetics. Hybrid materials have beendeveloped to preserve the bulk properties of traditional polymers whilemaking their molecular chains look more like proteins. The elusive goalof molecular recognition in synthetic polymer systems has been reachedin certain cases. Polyacrylic gels have been designed as withrecognition capabilities by incorporating non-covalently crosslinkedantibodies. These proteins couple the reversible swelling character ofthe networks with molecular recognition by only swelling in the presenceof a specific antigen. The advantage of using synthetic polymericmaterials based solely on proteins or peptides is the high degree ofcontrol over properties. Peptides and proteins can be coded for specificproperties using a basic knowledge of inter and intrachain interactions.The present and future of biomedical materials development requires adegree of control prediction in design, synthesis and function of nextgeneration materials. Recent work with this principle in mind hasresulted in protein-based materials with properties analogous to morewidely used polymers as well as new properties. These new materials havebeen generated with a variable degree of efficiency and complexity

The development of drug delivery vehicles requires systems that respondto a specific cue in the biological environment before the release of adrug payload. This is also coupled with the desire for such new devicesto otherwise maintain structural integrity and avoid clearance from thebody. We have described sensitive gels with stimuli-sensitiverecognition very similar to recognition in proteins. By outlining theprinciples developed by analyzing theoretical mechanics ofheteropolymers, the underlying memory of macromolecule conformation isdiscovered and empirically verified. Essentially, their design includespolymerizing in the presence of target molecule, functional monomer,thermo-sensitive monomer, and end shielded post-crosslinking monomer.Some of these adsorption sites were destroyed upon gel swelling andreformed upon shrinking. Important contributions have been madedescribing the nature of recognition in low cross-linked systems, and itis only a matter of time when intelligent gels can recognize other typesof molecules.

The present invention includes imprinted gels or chains possessingcertain macromolecular architecture with binding abilities could be usedas the sensing elements within analyte sensitive controlled releasesystems. Analyte sensitive polymer networks have been the focus of muchresearch (mostly saccharide recognition) and have been designed in anumber of ways.

Balancing pharmaceutical research for new drugs to treat human illnessand disease with economic factors to minimize the cost of drug therapyhas led to controlled and targeted drug delivery products. The goal ofcontrolled drug delivery is to reduce the cost of treatment by allowingsmaller, yet equally effective, dosages through a regulating device.Some drugs have very short half-lives in the human body, and large dosesof these drugs are metabolized rapidly, while other drugs, such as manyof the new protein drugs, are very fragile in the harsh environmentinside the body. Controlled release devices can prolong the release timefor the former, allowing effective dosages, and can protect sensitivedrugs until the point where they are to be delivered.

In the past, drug delivery devices have been limited to systems such astablets, capsules, powders, droplets, ointments and injections. Suchsystems while useful in treating some diseases have certaindisadvantages: (1) they are difficult to regulate drug delivery; (2)they deliver their bioactive agent (drug) relatively fast; and (3) agentdelivery is usually decreasing with time

It must be noted that although the description above uses a drug as anactive agent, similar problems have been observed with release ofdelivery of other active or bioactive agents, such as (but not limitedto): pesticides, herbicides, other agricultural products, molluscicides,other marine biology products, agents that kill ticks, flees, etc.,essential oils, perfumes, agents used in kitchen products, whiteningagents used in laundry detergents.

More recently, systems have been developed which allow controlledrelease of drugs to targeted areas of the body. Some methods used forthe controlled delivery of drugs include: inserts and implants,transdermal systems, oral delivery systems, nasal delivery systems,vaginal delivery systems, rectal delivery systems, ocular deliverysystems and bioadhesive/mucoadhesive systems.

Most of these systems while solving the problem of prolonged delivery ofactive agents, are not as efficacious in applications when the patientis unwilling or unable to take the necessary drug (payload) at aspecific time or specific interval. More precisely, such systems cannotcontrol the problem of patient compliance, a significant problem in thisindustry.

The use of carriers sensitive to the surrounding environment, such aspH-sensitive or temperature-sensitive systems have been reported in thefield. Indeed, investigators have reported methods of delivering drugs,active agents and bioactive agents in response to changes in pH ortemperature of the surrounding fluid.

Clearly, such systems can improve the pattern of delivery by beingtriggered to release their payload when a particular pH prevails in thesurrounding fluid. For example, numerous drug delivery systems have beenpatented where the passage from the stomach (low pH) to the upper smallintestine (high pH) triggers the release of an active compound (drug).Often, such systems are accompanied by selective targeting to varioustissue sites. For example, the so-called mucoadhesive drug deliverysystems are based on polymeric materials which adhere to the mucin layerof a biological membrane for some length of time. The desired drug isloaded into the polymers. Once introduced to the body, the polymercarrier begins to swell, allowing the release of the drug. Because thepolymer binds to the mucin layer of the membrane, the drug is releasedlocally and is thought to be able to absorb more easily across themembrane into the bloodstream. Some possible routes of administrationfor mucoadhesive systems include: the nasal, ocular, buccal,gastrointestinal, vaginal and rectal areas.

There are several distinct advantages in using controlled releasesystems over other methods of drug delivery. First, the drug can bedelivered at a relatively constant concentration. Thus, the drugconcentration can be maintained at a level that is higher than thetherapeutic level of the drug, but lower than the toxic level. In thecase of tablets, the drug concentration steadily increases until all ofthe drug has been released. At this point the concentration of drug inthe body may be above its toxic level. Once the drug has been releasedfrom the tablet the concentration decreases until a subsequent dose istaken. A second advantage of this type of drug delivery is that the rateand time period of delivery can be controlled depending on theproperties of the polymer system.

However, the previous systems do not possess the additional advantage ofintelligence of recognition of not just a change in pH or temperature,but in response to a finite concentration of an external analyte, acompound with special desirable or undesirable properties.

Most if not all of these systems, whether passive of pH- or T-sensitivehave a structure that belongs to the category of hydrogels. Hydrogelsare highly biocompatible which makes them appropriate for a number ofpharmaceutical and medical (but also cosmetic, food and consumer)applications. In addition to drug delivery carriers, hydrogels arebiomaterials used as contact lenses and scaffolds for tissue engineeringapplications to name only a few of the potential roles. The polymernetwork can contain homopolymers or copolymers with the chemicalstructure determining the properties of the hydrogel.

The network structure of the hydrogel can be characterized by a numberof parameters. Three parameters that I will discuss here are the polymervolume fraction in the swollen state, v_(2,s), the molecular weightbetween crosslinks, M_(c) and the distance between crosslinks also knownas the mesh size. The values for these parameters can be determinedempirically or by theoretical calculations.

The polymer fraction in the swollen state is a measure of how much waterthe hydrogel can imbibe when placed in an aqueous environment. Theability of hydrogels to retain large amounts of water makes them similarto natural tissue and may contribute to their high biocompatibility.Both the molecular weight and distance between crosslinks give anindication of how highly crosslinked the network is. Due to therandomness involved with polymer formation, these parameters can only begiven as average values throughout the hydrogel. These parameters canindicate how much space is available for diffusion in and out of thehydrogel. This value, along with the size of the agent to be delivered,will be important in determining the release kinetics of the agent fromthe hydrogel in drug delivery applications. The degree of swellingpresent in the network will affect the mesh size and therefore aphysiologically-responsive hydrogel that swells when presented withcertain stimuli can have different release kinetics at different sitesin the body.

pH-Responsive hydrogels are composed of ionic networks and swell inresponse to pH changes. This swelling behavior is controlled by theionization of the pendant groups in the network. Charged groups exhibitelectrostatic repulsion that leads to imbibition of water and increasedmesh size. This event also depends on the level of crosslinking presentin the hydrogel. Highly crosslinked materials will not be able to swellto as high a degree as materials with lower crosslinking ratios due todecreased chain mobility. The degree to which a hydrogel network willswell is also dependent upon the ability to imbibe water. Hydrogels withhydrophilic groups can imbibe more water than those with hydrophobicgroups and can therefore swell to a greater extent. Thehydrophobicity/hydrophilicity of the network will therefore also have animpact on the diffusion of any compound embedded within a hydrogelnetwork. An example of a monomer that will create an ionic hydrogel withpH-responsive swelling is methacrylic acid (MAA). When the pH of theenvironment is greater than the pK_(a) of the carboxylic acid groups inMAA, they become ionized and cause interchain repulsion. The pK_(a) ofthis group in poly (methacrylic acid) is approximately 4.9 making the pHshift from the stomach to upper small intestine (1.5-6) appropriate tochange the ionization of the carboxyl groups. The charged groups arealso hydrophilic and allow water to enter the network and continue theswelling process.

The process of ionization is reversible depending on the pH of theenvironment. If the MAA is grafted with another polymer capable offorming hydrogen bonds, like poly(ethylene glycol) (PEG), then hydrogenbonds can form between the chains when in the protonated state at lowpH. This pH-dependant formation of hydrogen bonds provides another meansby which the network exists in a compact state at low pH and a more openstate at the elevated pH. Hydrogels that exhibit this activity aretermed pH-responsive complexation hydrogels.

Through the use of monomers with side chains containing groups withpK_(a) values in the range desired, a pH-responsive hydrogel could bedesigned much in the same manner as enteric coatings. Hydrogels canexhibit swelling to different degrees based on the intensity of thestimulus and this could be used to target release of multiple compoundsat different sites. For example, if a hydrogel swelled and increased itmesh size sufficiently to release a small compound at one pH and showedincreased swelling at a pH later in the gastrointestinal tract, like thecolon as opposed to the small intestine, it could release a secondlarger agent at this location. The variability of the hydrogel deliverysystem in what it will respond to and how it will respond makes it anattractive candidate for numerous clinical applications includingtargeted drug delivery.

These hydrogels can be used for delivery of a variety of therapeuticagents. For example, previous work in our laboratory has focused on theuse of hydrophilic polymer carriers for oral delivery of proteins suchas insulin. The loading of proteins into the hydrogels was done byimbibition, where the polymer is swollen in a solution containing theprotein and collapsed at low pH to trap the protein inside.

Recognitive Materials—Molecular Recognition. The recognition of aspecific molecule out of a whole host of competing species is essentialto all life processes. It is this ability that allows for the properfunctioning of enzymes, antibodies, receptors, and signaling molecules.Ultimately, the design of biomaterials will include this molecularrecognition ability, whether it is a smart system that recognizes onlydiseased cells, an implantable device with a tailored surface that doesnot elicit an immune response, or a sensor that can track levels of aspecific compound in situ. In addition to use as biomaterials, thecreation of synthetic materials with recognitive abilities will havegreat benefits in the areas of separations, assays, catalysis, and masstransport.

Synthetic Systems for Molecular Recognition. Undoubtedly, methods forthe creation of materials with recognitive abilities similar to thoseshown in biological molecules such as enzymes and antibodies have beenheavily sought after.

Molecular Recognition with Crosslinked Networks. By crosslinking thepolymer chains, it is possible to restrict the number of conformations agiven chain may adopt. In the formation of a configurationallybiomimetic imprinted polymer (CBIP), interactions between a templatemolecule and the monomer feed molecules leads to the creation of abinding site that is subsequently locked in by polymerization andcrosslinking. To date, imprinted structures have been successfully usedin chromatographic applications [34-39], as sensors [40-42], and even ascatalytic elements [43-46].

Wulff, et al. [34] demonstrated the first simple molecularly imprintedpolymers (MIPs) utilizing monomers that had been covalently linked tothe functional monomers in order to establish a proper 1:1stoichiometric ratio. After polymerization, the template molecule wasfreed through lysis. This covalent technique of imprinting has severaladvantages including efficient use of all available functional groupsand a propensity to form a more uniform binding pocket, but isrestrictive in what monomers may be used. A technique later pioneered inthe laboratories of Mosbach involves imprinting a freely solubletemplate without the use of covalent linkages [47, 48]. This techniqueallows for greater flexibility in the choice of functional monomers aswell as template molecule. However, reaction conditions need to be morestrictly controlled to maximize interaction between template and monomermolecules. In addition, a number of different binding sites may formleading to some nonspecific binding.

The molecular imprinting procedure. The production of a successfullyimprinted polymer results in a material with recognitive properties.While many polymerization techniques are amenable to the imprintingprocedure, most utilize a free radical technique with either thermal,ultraviolet, or redox methods providing the initiating radicals [49].Common monomers include the methacrylate and acrylate family ofmolecules, acrylamides, and other vinyl derivatives as these are readilyavailable, polymerize easily with the free radical technique, and areavailable with a number of functional groups [50]. In addition tomonomer molecules with an array of functional groups, it is crucial tohave crosslinking agents that incorporate well into the polymerization.Usually, a crosslinking agent is selected such that it has similarreactivity to the monomers used so as to form a network uniform incrosslinking density [51].

The prepolymerization mixture includes the functional monomers,crosslinking agent, initiating species, template, and solvent ifdesired. As free radical polymerizations are sensitive to the presenceof radical scavengers such as dissolved oxygen, the mixture is firstpurged with an inert gas such as nitrogen. Polymerization is theninitiated. As suggested by Pande et al. [28] in their work on randomheteropolymers, successful imprinting is more likely to occur at lowertemperatures since entropic effects are lessened, which suggests the useof either a redox or UV initiation method. However, many groups havesuccessfully used thermal initiation, albeit at lower temperatures thannormally seen for thermal polymerizations.

Following polymerization, the imprinted polymer is swollen in solvent tofacilitate the removal of the template molecule. Often times beforedialysis, the crosslinked material is crushed and sieved to produceparticles of a given diameter in order to facilitate mass transfer. Oncefree of template, the MIPs are subjected to analysis of their bindingability through a variety of techniques including liquid chromatography,NMR, and microcalorimetry.

Yu and Mosbach [35] studied the influence of crosslinking density on therecognitive ability of a non-covalently formed MIP imprinted for theseparation of Boc-D-Trp and Boc-L-Trp enantiomers. In their studies, itwas shown that the ability to separate enantiomers decreases as thecrosslinking density is decreased. This supports work done by Wulff, etal. [52] with covalently imprinted materials where in addition to adecrease in recognition, there was also a minimum amount of crosslinkingneeded for recognition. While increases in crosslinking density arefavorable to the recognitive ability of a MIP, the resultant decrease innetwork mesh size may act to hinder diffusion through the network,especially limiting for the recognition of large molecules such asproteins.

Applications of molecularly imprinted materials. Historically, the mainapplication of MIPs has been in chromatographic uses. Crosslinkedrecognitive materials are formed with through the imprinting process andare then crushed into particles suitable for packing intochromatographic columns [49], or used in thin layer chromatography [37].The recognitive ability of the particles allows for affinitychromatography whereby only the compound of interest is bound by thecolumn and all other species elute freely. This has worked especiallywell for the separation of enantiomers, a classically difficultseparation. In 1974, Wulff, et al., [34] first suggested the formationof configurationally biomimetic imprinted polymers for the separation ofD,L-glyceric acid. Since then, a significant amount of work has beenfocused on creating MIPs for separations, including enantiomers ofTryptophan derivatives by Yu and Mosbach [35], nicotine-like compoundsby Andersson et al. [36], D- and L-phenylalinine anilide by Kriz et al.[37], penicllin G and related compounds by Cederfur et al.[38], andamino acids by Kempe and Mosbach [39].

More recently, MIPs have been employed as sensing elements due to theirmolecular recognition abilities. Due to the minute amounts typicallybound, a highly sensitive quartz crystal microbalance (QCM) is used todetermine the mass of analyte absorbed. Detection by QCM has been usedby Haupt et al. [40] for the detection of R- and S-propanololhydrochlorides, by Liang et al [41] for the detection of epinephrine,and by Hilt et al. [42] for the detection of D-glucose.

Traditionally, the design of biomaterials has focused onbiocompatibility—the propensity for a material to not invoke a foreignbody response upon implantation or contact in the body. Numerous studieshave been done to tailor surface properties so as to not elicit animmune response. The main method has been to functionalize the surfacewith a hydrophilic molecule, such as grafted poly(ethylene glycol)chains, to mask the foreign surface from protein adsorption. However, itis now being recognized that molecular recognition may play an importantrole in future biomaterials design. Materials that show goodbiocompatibility are being further enhanced to include molecularrecognition. Articles by Ratner [53] and by Peppas and Langer [54]discuss how building recognitive abilities into biomaterials has thepotential to radically advance the field. Potential applicationsinclude: (1) materials that invoke healing pathways to rebuild tissue inthe implantation area; (2) combined sensing element/controlled releasedevice to meter and release appropriate amounts of therapeuticcompounds; (3) recognitive materials specific to toxins or deleterioussignaling molecules (such as angiotensin) for rapid detoxification inthe blood stream; and (4) antibody or enzyme mimics for in vivo use fromsynthetic materials.

Researchers working towards these goals have focused on the creation ofmaterials that are suitable for use in the body yet interact with thebiomolecule of interest. Byrne et al. [55, 56] and used an imprintingtechnique to create hydrogels capable of binding D-glucose. Thistechnique was later coupled with sensing technologies developed by Hiltet al. [57] to form a microsensor capable of detecting D-glucose [42].Other glucose responsive materials have been formed by Oral and Peppas[58] utilizing star polymers and by Parmpi and Kofinas [59]. In additionto small biomolecules, Bolisay et al. prepared a configurationallybiomimetic imprinted material suitable for the detection and screeningof baculoviruses [60]. Molecular imprinting is well-known and can beconducted using one or more biomolecules, e.g., Acetaldehyde (metabolismbyproduct); Adenine, adenosine 5V-triphosphate (ATP); Amino acid andpeptide derivatives: Z-L-Tyr-OH; Z-L-Phe-OH; Z-DL-Phe-OH; Z-L-Glu-OH;Boc-L-Phe-Gly-Oet; Z-L-Ala-L-Ala-OMe; Z-L-Ala-Gly-L-Phe-OMe; Z-L-Phe-OH;Ampicillin (penicillin antibiotic); a-Amylase (enzyme); Angiotensin II(SA) (competitive inhibitor of, peptide hormone angiotensin II);Bupivacaine (anaesthetic drug); Butein (active anti-EGFR inhibitor);Caffeine (stimulant drug); Cephalexin (antibiotic drug; ina-aminocephalosporins class); Chlorphenamine (anti-histamine drug);Clenbuterol (h adrenergic blocker); Cortisone (steroid); Creatine(metabolite); Creatinine (metabolite); Cholesterol (steroid); Cholicacid sodium salt (bile acid); Carbohydrates: glucose; lactose, maltose,glucose; Glucose; Maltose; lactose; cellobiose; Carbohydratederivatives: octyl-glucoside; p-nitrophenyl fucoside, p-nitrophenylgalactoside; Peracetylated phenyl a- and h-D-galactosides; Diazepam(i.e., valium; benzodiazepine anxiolytic drug); Enkephalin(neuropeptide); Ephedrine (stimulant drug); Epinephrine (adrenalinehormone); Estradiol (estrogenic steroid hormone); Ethynylestradiol(estrogenic steroid hormone derivative); 9-ethyladenine (nucleotide basederivative); 9-ethyladenine acetate (nucleotide base derivative);Glucose oxidase (enzyme); L-glutamine (amino acid); Histidine(N-terminal) dipeptides; Homocysteine (non-essential amino acid);Horseradish peroxidase (enzyme); Ibuprofen (non-steroidalanti-inflammatory drug); Ketoprofen (non-steroidal anti-inflammatorydrug); Lysozyme (enzyme); Morphine (narcotic analgesic drug); Naproxen(non-steroidal anti-inflammatory drug); Nerve agent degradationproducts; (S)-nilvadipine (dihydropyridine calcium antagonists);Nucleoside base derivatives: tri-O-acetyl adenosine; tri-O-acetylguanosine; di-O-acetyl thymidine; tri-O-acetyl cytidine; tri-O-acetyluridine; Nucleotide base derivatives: 9-ethyladenine; 1-propyl thymine;1-propyl cytosine; 1-cyclohexyl uracil; Oxytocin (hormone); Paracetamol(i.e., acetaminophen, analgesic); Phenylalanine (amino acid);(E)-piceatannol (active anti-EGFR inhibitor); Propanolol (h adrenergicantagonist); Quercetin (active anti-EGFR inhibitor); Ribonuclease A(enzyme); Ricin A and B Chains (toxin bean lectin); (S)-ropivacaine(anaesthetic); Scopolamine (anti-cholinergic, anti-infective; andanalgesic alkaloid drug); Sulfonamides (antibiotic drug); Testosterone(steroid hormone); Tetracycline (antibiotic drug); Theophylline(Bronchodilator drug); Timolol (h adrenergic blocker); Trypsin (enzyme);Tyrosine (amino acid); Tyr-Pro-Leu-Gly-NH2 (tetrapeptide);Leu-enkephalin; Leu-enkephalin; Morphine; Morphine; Ampicillin;S-propranolol; D-phenylalanine; Adenine; 9-ethyladenine; 9-ethyladenine;9-ethyladenine acetate; Cholesterol; Homocysteine; Trypsin;Theophylline; see, e.g., Hilt & Byne, Configurational biomimesis in drugdelivery: molecular imprinting of biologically significant molecule,Advanced Drug Delivery Reviews 56 (2004) 1599-1620, relevant portionsand citations incorporated herein by reference.

Protein Imprinting. The potential applications for a recognitivematerial capable of binding a protein are numerous, including diagnosticdevices for protein assays, systems for use in immunochemistry, andseparation media for extremely complicated protein mixtures. Productionof such materials, however, is difficult for several reasons. First, itis known that the presence of water reduces the interactions betweentemplate and monomer since the water molecules compete for hydrogenbonds [33]. Most imprinting, therefore, is done in the presence ofnon-aqueous media. However, peptides and proteins are especiallysensitive to differing solvent conditions and may denature in harshsolvents. Secondly, the large diameter of protein molecules may precludethe use of a densely crosslinked polymeric network since the mesh sizeof the network is too small to allow for efficient diffusion. It is alsounclear how selective a protein imprinted material can be made, andwhether subtle changes, such as the process of site directedmutagenesis, can be differentiated by these materials.

To date, several attempts have been made at creating MIPs for therecognition of oligopeptides and proteins. Shnek et al. [61] and Kempeet al. [62] focused on a surface imprinting approach whereby monomerscapable of complexing a metal ion were polymerized into a MIP for withan affinity for proteins with surface exposed histidine residues.Polymers imprinted in the bulk have been prepared for the recognition ofpeptides by Andersson et al [63] and for the recognition of proteins byVenton and Gudipati [64]. Recently, Rachkov and Minoura [65] havedemonstrated a method where only a fragment of the protein is used inthe imprinting process, leading to a site that is favorable for aportion of the whole protein. This so called “epitope approach” mimicsthe ability of antibody molecules to recognize a portion of a proteinstructure.

Essential oils. Essential oils are complex mixtures of numerouscompounds, they are aromatic oily liquid and can be extracted fromvarious parts of the plants. Some of the main chemical groups found inessential oils include alcohols, aldehydes, esters, ethers, ketones,phenols and terpenes. Their use is mostly related to food as flavoring,to perfumes as fragrances and to pharmaceuticals for their functionalproperties. To date they are widely used as air freshener, severalpatents are reported dealing with their applications as deodorant, someare related to their use as good smelling insect repellent. As reportedin the review proposed by Burt (2004), antimicrobial properties of someessential oils have long been recognized; in particular they have beenshown to exhibit antiviral, antimycotic, antitoxigenic, antiparasiticand insecticidal properties. In the literature several works reports ontheir use to improve the shelf-life and safety of food and packagedfood.

Controlled release systems. As reported by Peppas and Am Ende (1997),the incorporation of fragrances in polymers for the purpose ofcontrolled release over a period of 12 or more hours has been studied.Incorporation of essential oils in polymers could lead to new andinnovative products for prolonged delivery of these compounds. Peppasand Brannon-Peppas (1996) provided a wide view on papers dealing withthe use of systems in which the fragrances are released from differentmatrices. In addition to those one, the release of linalool and linalylacetate (the major components of aromatic lavender essential oils) frombiopolymer gliadin-based nanoparticles (Duclairoir et al., 2002), therelease of eucalyptus essential oil from alginate complex capsule (Changand Dobashi, 2003) and the release of limonene from polysaccharidesmatrices (Secourad et al., 2003) were studied. Besides, Nakayama et al.(2003) studied the release of orange essential oils from temperatureresponsive membranes obtained by UV polymerization of mixture ofN-isopropyl acrylamide and Polyethyleneglycole dimethacrylate. Thesereferences provide interesting information on the fragrance releasemechanism but, due to the requirements and characteristics of essentialoils, swelling-controlled release systems are highly desirable devicesfor such applications, relevant portions incorporated herein byreference.

The preparation of the above mentioned devices requires theunderstanding of the thermodynamic of the three-components system,consisting of the polymer, the fragrance and the liquid which is usuallyin contact with the device. The release of a fragrance initiallyembedded into the polymer matrix into the surrounding solvent is limitedby several factors: the initial amount of fragrance loaded into thepolymer, the solubility of the fragrance in the solvent, the equilibriumpartition coefficient of the fragrance between polymer and solvent andthe diffusional barrier.

The present disclosure generally relates to biomimetic polymer networkcompositions, methods of forming such polymer compositions, and methodsof using such compositions. These compositions and have improvedproperties that make them useful for a variety of applications; inparticular, the loading and delivery of therapeutic agents.

The biomimetic polymer networks of the present disclosure generallyinclude a polymer network having architectures that have selectiveaffinity for a moiety. Such biomimetic polymer networks may have shapespecific cavities that match the moiety, as well as chemical groupsoriented to form multiple complexation points with the moiety. In termsof selectivity, the resulting polymer networks are selective due to theparticular chemistry of the binding site, the orientation andstabilization of the chemistry in a crosslinked matrix, as well as bythe size and shape of the site for the template biomolecule.

In some embodiments, the biomimetic polymer networks may furthercomprise a moiety. Such compositions may be capable of releasing themoiety in a relatively controlled fashion. The moiety may be present ona target compound, for example, a therapeutic agent. Accordingly, thecompositions and methods of the present disclosure may be used in thetreatment of a disease. For example, the compositions of the presentdisclosure may be used as a vehicle to deliver a therapeutic agent to asubject (e.g., a human) in need thereof. The compositions of the presentdisclosure also may be used to form a medical device or an article. Thepresent disclosure also provides methods of forming a biomimetic polymernetwork of the present disclosure.

The moiety may be any portion of a molecule recognized by a biomimeticpolymer network of the present disclosure. The moiety may be covalentlybound to a target compound, for example, a therapeutic agent. In thisway, the moiety may be used to associate a target compound with abiomimetic polymer network of the present disclosure. The moiety shouldeither already be present on the target compound or capable of beingconjugated to a target compound. Conjugation of moieties to therapeuticagents is known in the art, for example, as disclosed in A. Wong and I.Toth, Curr. Med. Chem. 8:1123-36 (2001), the relevant disclosure ofwhich is incorporated by reference. Examples of suitable moietiesinclude, but are not limited, to sugars (e.g., glucose), carbohydrates,peptides, and functional groups. A specific example of a therapeuticagent that comprises a moiety is streptozotocin (R. R. Herr, et al., J.Am. Chem. Soc. 89:4808-09 (1967)), which has a glucose moiety.

In certain embodiments, the moiety is a sugar. For example, the sugarmay be a monosaccharide. Monosaccharides have the chemical formula(CH2O)n and the chemical structure H(CHOH)nC═O(CHOH)mH. If n or m iszero, it is an aldose, otherwise it is a ketose. Monosaccharides mayinclude aldoses, trioses (e.g., glyceraldehyde), tetroses (e.g.,threose), pentoses (e.g., ribose, xylose), hexoses (e.g. glucose,fructose, mannose, galactose), ketoses, trioses, tetroses, pentoses(e.g., ribulose), hexoses (e.g., fructose). Any of the L and D isomersof a sugar also may be used, although the D isomer may be more preferredfor biological applications. Other examples of suitable sugars includepolysaccharides. Polysaccharides have a general formula ofC_(n)(H₂O)_(n-1) where n is usually a large number up to 500.Disaccharides, such as, for example, sucrose, lactose, maltose, and thelike may be used. Yet another example of suitable sugars includesoligosaccharides and low molecular weight carbohydrates (e.g., having amolecular weight no greater than about 2,000 Da). Further, each carbonatom that supports a —OH group (except for the first and last) ischiral, giving rise to a number of isomeric forms all with the samechemical formula.

Specific embodiments may use the following monosaccharides as moieties:monoses, dioses, trioses, tetroses, pentoses, aldo-pentoses, includingarabinose, ribose, deoxyribose and xylose, keto-pentoses includingribulose, hexoses including aldo-hexoses such as: allose, altrose,galactose, glucose, mannose and talose, and keto-hexoses such asfructose, heptoses including keto-heptoses such as mannoheptulose andsedoheptulose, octoses such as octolose,2-keto-3-deoxy-manno-octonateand and nonoses such as sialic acid.

Specific embodiments may use mucopolysaccharides. Mucopolysaccharidesare long unbranched polysaccharides consisting of a repeatingdisaccharide unit. This unit consists of an N-acetyl-hexosamine and ahexose or hexuronic acid, either or both of which may be sulfated.Members of this family vary in the type of hexosamine, hexose orhexuronic acid unit they contain e.g. glucuronic acid, iduronic acid,galactose, galactosamine, and glucosamine. They also vary in thegeometry of the glycosidic linkage. Specific example polysaccharidesthat may be used as moieties include: chondroitin sulphate, dermatansulphate, keratan sulphate, heparan sulphate, heparin, sodium heparin,hyaluronic acid and hyaluronan.

In other embodiments, the moiety may be a lipid or a short amino acidsequence (e.g., a sequence of about twenty amino acids in length). Inparticular, lectins may be used as a moiety. Lectins arecarbohydrate-binding proteins involved in a variety of recognitionprocesses and exhibit considerable structural diversity. A largevariability in quaternary association resulting from small alterationsin essentially the same tertiary structure is a property exhibitedspecially by legume lectins. The strategies used by lectins to generatecarbohydrate specificity include the extensive use of water bridges,post-translational modification and oligomerization. Othercarbohydrate-based structures may be used as moieties may be located athttp://www.chem.qmul.ac.uk/iupac/2carb/ (accessed Apr. 27, 2006),incorporated by reference herein.

In general, the compositions of the present disclosure have enhancedaffinities (e.g., impart greater affinity, bound ratios greater than 1)for a chosen moiety, among other things, allowing for increased loadingefficiency. Accordingly, the compositions of the present disclosure alsomay be used to increase the loading of a target compound or control therelease rate of a target compound or both. The compositions of thepresent disclosure also may be used for delivery of a therapeutic agent.For example, the compositions of the present disclosure may be used asan excipient or as a vehicle for a therapeutic agent. Specifically,higher quantities of a therapeutic agent having a moiety can be loadedwithin the biomimetic polymer networks of the present disclosure,therefore enabling for higher doses to be loaded. The release of amoiety may be tailored to give a desired release profile, for example,for sustained release of a therapeutic agent. Thus, when the moiety isbound to a therapeutic agent, treatment with the therapeutic agent maybe optimized.

The compositions of the present disclosure may be formed usingconfigurational biomimetic imprinting. Configuration biomimeticimprinting techniques generally involve forming a prepolymerizationcomplex between the template molecule (e.g., a moiety) and functionalmonomers or functional oligomers (or polymers) with specific chemicalstructures designed to interact with the template either by covalentchemistry or noncovalent chemistry (self-assembly) or both. Once theprepolymerization complex is formed, the polymerization reaction occursin the presence of a crosslinking monomer and an appropriate solvent,which controls the overall polymer morphology and macroporous structure.Once the template is removed, the product is a heteropolymer networkwith specific recognition elements for the template molecule.

The network structure depends upon the type of monomer chemistry (i.e.,anionic, cationic, neutral, amphiphilic), the association strength andnumber of interactions between the monomers and template molecule, theassociation interactions between monomers and pendent groups, thesolvent type and the amount of solvent in the mixture, the reactivityratios of the monomers, and the relative amounts of reacted monomerspecies in the structure. Since noncovalent forces are weaker thancovalent bonds, an increased number of interactions are needed forstable binding and recognition. On a per-bond basis, noncovalent bondsare 1-3 orders of magnitude weaker. Therefore, a greater number ofnoncovalent bonding with matching structural orientation is needed foraqueous recognition.

A wide variety of polymers may be used to form the heteropolymernetwork. These include polymers produced by reaction of acrylamides andall their substituted structures including: methacrylamide,ethacrylamide, isopropyl acrylamide, etc., acrylic acid, methacrylicacid, ethacrylic acid, all alkyl acrylic acids, any dicarboxylic acid,such as crotonic acid, phthalic and terephthalic acid any tricarboxylicacid with itself another monomer of the above list (forming acopolymer), two other monomers from the above list (formingterpolymers), or three or more monomers from the above list forminghigher order coploymers. The above may be in linear, branched or graftedform, the grafted chains being exclusively one polymer or copolymers ofthe above, ionically bound or complexed by hydrogen bonds.

The above may be crosslinked in the presence of crosslinking agents toform insoluble but swellable gels or networks, having the ability toabsorb water, physiological fluids, buffers or salt solutions with finalswelling as low as 1 weight % of water and as high as 99.9% water.

The above crosslinking may be achieved with ethylene glycoldimethacrylate, ethylene glycol diacrylate, ethylene glycoltrimethacrylate, ethylene glycol diacrylate, ethylene glycol multimethacrylate where “multi” stands for n=4 to 200 units ethylene glycolmulti acrylate where “multi” stands for n=4 to 200 units same as abovebut propylene glycol multi methacrylate where “multi” stands for n=1 to200 units same as above but alkylene glycol multi methacrylate where“multi” stands for n=1 to 200 units. One may also use higher orderacrylates and methacrylates including but not limited to 1,1,1trimethylolethane trimethacrylate (TrMETrMA, Molecular Weight 324.4);1,1,1 trimethylolpropane triacrylate (TrMPTrA, Molecular Weight 296.3);1,1,1 trimethylolpropane trimethacrylate (TrMPTrMA, Molecular Weight338.4); pentaerythritol triacrylate (PETrA, Molecular Weight 298.3);glycerol propoxy triacrylate (GlyPTrA, Molecular Weight 428.5);pentaerythritol tetraacrylate (PETeA, Molecular Weight 353.2);ethoxylated 1,1,1 trimethylolpropane triacrylate (ETrMPTrA, MolecularWeight 428); glycerol propoxylated triacrylate (GlyPTrA, MolecularWeight 428) and glycerol trimethacrylate (GlyTrMA, Molecular Weight396.3). One may also use with “star polymers” or “dendrimers” with up to72 independent chains ending in acrylates or methacrylates.

The initiator may be IRGACURE® products of the Ciba Geigy companyincluding IRGACURE 184, IRGACURE® 379, CIBA® IRGACURE® 819, and CIBA®IRGACUREL® 250. Any other photoinitiator may also be used. The initiatormay also be any peroxide including but not limited to benzoyl peroxide,cumyl peroxide, etc. or Azobis isobutyronitrile.

In some embodiments, the biomimetic polymer network of the presentdisclosure may be formed using a template molecule (e.g., D-glucose) andfunctional monomers selected to match corresponding template molecule(e.g., glucose binding protein residues, such as aspartate, glutamate,and asparagines, as well as biological mechanisms of action that involverecognition The template molecule may be added in stoichiometric amountsin regard to the functionality of the template molecule. Since solventinteraction can stabilize or destabilize binding in noncovalent systems,functional monomers may be selected based on optimizing specificnoncovalent, self-assembly interactions (hydrogen bonding) with thetemplate molecule within an aprotic solvent (e.g., dimethylsulfoxide).Such techniques are generally applicable to template molecules, in whichhydrogen bonding, hydrophobic, or ionic contributions will directrecognition of the moiety. The formation of an exemplary biomimeticpolymer network of the present disclosure according to the methods ofthe present disclosure is described below.

The multilayered mimetic structures may be constructed from a variety ofcoating processes, including pan coating, air-suspension coating,centrifugal extrusion, vibrational nozzle coating, supercritical fluid(SCF) based processing, fluidization (both conventional Wurster coaterssuch as the Glatt device) and rotating, or spray-drying.

The multilayered mimetic structures shown in FIGS. 1-2 may beconstructed using a Grow Max spouted bed assisted with a draft tube anda bottom spray, also known as the Wurster configuration. The height ofthe bed is 535 mm and the diameter of the bed is 160 mm at the bottomand 300 mm at the top.

The bed is equipped with a sampling port to allow the microcapsule to bewithdrawn and analyzed during the coating process. The bed is alsoequipped with a viewing window to allow the observation of thefluidizing microcapsule during the coating process. The length of thedraft tube to be used is preferably 200 mm and is located 15 mm abovethe air distributor. The diameter of the draft tube is 70 mm. Thepneumatic spray nozzle with a liquid caliber of 1.0 mm can be used toatomize the dispersion fed to the bed. A bag filter with 5 μm openingcan be used to prevent the microcapsules from escaping through the topof the bed. The bag filter must be shaken frequently to prevent themicrocapsule from adhering to the bag filter. The schematic diagram ofthe Wurster bed apparatus is shown in FIG. 2.

The advantages of the Wurster process compared to conventional fluidizedbed process are: (i) the high inertia introduced in the Wurster processto circulate the particles prevents the agglomeration of the particlesand (ii) the recycling profile creates a uniform coat around theparticles. The circulating pattern of the particles inside a Wurster bedis made possible by a draft tube. The air inlet is concentrated justunder the draft tube. This creates a high air velocity inside the drafttube and a vacuum like effect at the base of the draft tube. Theparticles collected under the bed are readily pulled by the air inletinto the draft tube and being carried to the top of the bed. At the exitof the draft tube, there is a sudden expansion, from the diameter of thedraft tube to the diameter of the chamber. This expansion causes theparticle to lose its velocity gradually and finally gravity will pullthe particles back down along the wall of the chamber to the base andthe process will be repeated.

Generally, a Wurster bed is equipped with a bottom spray, and hence,every time the particles pass through the based of the draft tube, latexdispersion will be sprayed on the particles. This process repeats everytime the particles pass through the bottom of the draft tube, andfinally, a uniform coating can be obtained. The ability to coat suchsmall particles uniformly has broadened the applications of the Wursterprocess for particulate drug delivery systems.

The Wurster coating process employed in the present disclosure is veryversatile and can be applied for many different materials. The processhas been shown to work for many different type of cores includingcalcium carbonate crystal, glass beads and lactose [1-6]. Many latexdispersions have also been shown to be able to be coated using thisprocess, such as poly(ethyl acrylate/methyl methacrylate/2-hydroxyethylmethacrylate) (P(EA/MMA/HEMA)), composite latex of core P(EAAMMA/HEMA)and P(NIPAAm) shell, crosslinked P(NIPAAm), Eudragit RS30D, L30D-55 andFS30D, and ethyl cellulose [1-6].

The Wurster coating process is a very harsh process. The processinvolves exposure to high temperature and high shear stress. The processrequires high temperature to dry the coated microcapsules. The highshear stress is required to atomize the latex dispersion before beingsprayed on the circulating microcapsules. Fortunately, the MIPnanoparticles have shown promising stability at high temperatures andhigh shear stresses.

One problem encountered with the coating process is the formation of a“snowman-like” structure due to agglomeration. These structures areproduced by the fusion of two microcapsules. This fused structurecreates weak spot along the contact point of the two microcapsules.Agglomeration prevents the formation of a single core microcapsule,which is the structure desired in this work. Several factors that affectthe degree of agglomeration using the Wurster bed include: the feed rateof the latex dispersion, the flow rate of the dried air to fluidize themicrocapsule and the temperature of the inlet air, the outlet air andthe bed and the like.

The network structure of the layers depends upon the type of monomerchemistry (i.e., anionic, cationic, neutral, amphiphilic), theassociation strength and number of interactions between the monomers andtemplate molecule, the association interactions between monomers andpendent groups, the solvent type and the amount of solvent in themixture, the reactivity ratios of the monomers, and the relative amountsof reacted monomer species in the structure. Since noncovalent forcesare weaker than covalent bonds, an increased number of interactions areneeded for stable binding and recognition. On a per-bond basis,noncovalent bonds are 1-3 orders of magnitude weaker. Therefore, agreater number of noncovalent bonding with matching structuralorientation is needed for aqueous recognition.

A wide variety of polymers may be used to form multilayered mimeticstructures. These include polymers produced by reaction of acrylamidesand all their substituted structures including: methacrylamide,ethacrylamide, isopropyl acrylamide, etc., acrylic acid, methacrylicacid, ethacrylic acid, all alkyl acrylic acids, any dicarboxylic acid,such as crotonic acid, phthalic and terephthalic acid any tricarboxylicacid with itself another monomer of the above list (forming acopolymer), two other monomers from the above list (formingterpolymers), or three or more monomers from the above list forminghigher order coploymers. The above may be in linear, branched or graftedform, the grafted chains being exclusively one polymer or copolymers ofthe above, ionically bound or complexed by hydrogen bonds. Forapplication in organisms, such as humans, the polymers that will havemost application are those that are biocompatible, and in some cases,also biodegradable.

The above may be crosslinked in the presence of crosslinking agents toform insoluble but swell-able gels or networks, having the ability toabsorb water, physiological fluids, buffers or salt solutions with finalswelling as low as 1 weight % of water and as high as 99.9% water.Typically, highly crosslinked nanoparticles with high T_(g) cannot fusetogether and form films because the mobility of the polymer chain ishighly restricted. This restriction on the mobility of the polymerchains prevents the polymer chains to entangle. The film formation ofthe hydrogel layer can be prevented by using highly crosslinkedparticles with high glass transition temperature, T_(g), (as is the casewith the MIP particles disclosed herein) and also by the addition ofhydrophilic linear polymer chains as spacers in between thenanoparticles. On the other hand, film formation is a necessity for theskin layer. Film formation improves the rigidity and integrity of theskin layer, and hence, provides the required barrier to prevent the drugtransport across the skin layer. The film formation in the skin layercan be enhanced by the addition of some plasticizers. The addition ofplasticizer to the MIP may turn out to be effective in lowering theglass transition temperature of the system, and hence, enhanced the filmformation of the coating layer.

Another problem with this multilayered mimetic system is the maximumsize. The size is controlled by the size of the syringe because themedical counter-part of this system is designed for injectable system.The size problem can be solved by mixing several differentmicrocapsules. For example, the size limitation would only allow themultilayered mimetic structures to have 16 layers, which would betranslated to 8 bursts. If a drug delivery system requires a burst everythree hours, then the system can only lasts for one day. However, if wemodify the multilayered mimetic structures, so that it would ruptureevery 12 hours and also the outer most layer is modified into fourdifferent batches; (i) one that will rupture after 3 hours; (ii) anotherthat will rupture after 6 hours; (iii) another after 9 hrs; and (iv)another after 12 hrs. By mixing these four different batches, the systemcan last for 4 instead of one day. This method of mixing made thesemultilayer mimetic structures to be very versatile and can easily betailored to fit any release profiles.

In order to show controllable properties, a mathematical expression ofthe system an be used to predict the burst process and when each drugrelease will occur. This expression is derived from the force balancebetween the inside and outside of the seal coat layer. The microcapsuleis assumed to be a perfect sphere with finite coating thickness. The MIPcoat has a tensile strength, which is dependent on the molecular weightof the material. The force balance can be simplified by dividing themicrocapsule into two equal hemisphere, and thus, canceling thex-component of the pressure. The pressure inside is larger than thepressure outside due to the osmotic pressure and the swelling of thehydrogel nanoparticles.

The y-component of this pressure will effectively act on the projectedarea of the hemisphere as represented by the left hand side ofEquation 1. The tensile strength of the seal coat layer on the otherhand will oppose this force along the rim microcapsule as represented bythe right side of Equation 1.

(P _(in) −P _(out))πr ²=τ(2πrl)   (1)

P_(out) and P_(in) are the pressures outside and inside of thehemisphere, respectively, τ is the tensile strength of the MIP coating,r is the radius of the hemisphere, and l is the thickness of the sealcoat layer. This equation assumes the radius of the hemisphere is muchlarger than the thickness of the MIP coat. Simplifying Equation (1), thetensile strength can be expressed as a function of the net pressure, theradius of the hemisphere, and the thickness of the MIP coat layer asshown in Equation (2).

$\begin{matrix}{\tau = \frac{\Delta \; \Pr}{2l}} & (2)\end{matrix}$

The tensile strength of polymers increases with increased crosslinkingand ultimately reaches an asymptotic value. This can be represented asEquation (3), where A and B are constants [7, 8]. The A and B constantsdepended on the degree of crosslinking and the intrapolymer interactionsof the polymer.

$\begin{matrix}{\tau = {A - \frac{B}{\overset{\_}{M_{n}}}}} & (3)\end{matrix}$

From Equations (3) and (2) we obtain the molecular weight (crosslinking)as a function of the net pressure, the radius of the hemisphere, thethickness of the MIP coat and the constants of the material as shown inEquation (4).

$\begin{matrix}{{\overset{\_}{M_{n}}(t)} = \frac{B}{A - \frac{\Delta \; \Pr}{2l}}} & (4)\end{matrix}$

Since the coating material of interest comes apart with swelling andparticle separation, M_(n) decreases with time. The swellingdisintegration of first order kinetics as described by Equation (5),where k is a rate constant.

M_(n) (t)= M_(n) e ^(−kt)   (5)

Inserting Equation (5) into Equation (4) and solving for t, we maycalculate the estimated lag time before the rupture process.

$\begin{matrix}{t = {- \frac{\ln\left\lbrack \frac{B}{{\overset{\_}{M_{n}}}_{i}\left( {A - \frac{\Delta \; \Pr}{2l}} \right)} \right\rbrack}{k}}} & (6)\end{matrix}$

This time parameter is an important parameter that can be tailored byseveral approaches: (1) select materials with different disintegrationkinetics resulted in different lag time by controlling the k parameter;(2) select materials with different degree of crosslinking andintrapolymer complexes thus affecting the lag time in term of the A andB parameters; (3) select material with high starting concentration andmolecular weight that can prolong the lag time; (4) increase thepressure difference between the inside and outside of the multilayeredmimetic structure by adding osmotic agents, such as salt and highmolecular weight hydrophilic polymers that could decrease the lag time;and, (5) increase the thickness of the seal coat layer could prolong thelag time.

There are several important assumptions in this system. The net pressuredifference between the inside and outside of the microcapsule is assumedto be constant throughout the degradation process. The net pressuredifference is assumed constant. The thickness of the coating layer isassumed to be constant throughout the process.

EXAMPLE 1

An example of the present invention was constructed for glucoserecognition followed by insulin release. The system was formed by usinga core, followed by a coating of the CBIP polymer in multiple layers.The core used was glass beads (size range 63-75 μm, Sigma, St Louis,Mo.) although potato starch or other materials such as calcium carbonatecrystals (size range 63-75 μm) could be used. The MIP or recognitivelayer consisted of a CBIP, hydroxy propyl cellulose (HPC) as a spacerand mannitol as a binder. Triacetin was used as a plasticizer to enhancethe film formation.

The MIP Hydrogel Coating. A CBIP coating was prepared by reacting 20 mlof acrylamide (Aam), 1.7 g of 2,2-dimethoxy-2-phenyl acetophenone (DMPA)as an initiator, 18 ml of dimethylsulfoxide (DMSO) as a solvent, and 8ml of ethylene glycol dimethacrylate (EGDMA) as a crosslinking agent.The system was reacted in the presence of 10 g D-glucose. The reactionwas carried out in a 100 ml glass reactor placed under a UV source(Dymax Ultraviolet Flood Cure System) and exposed to UV light with anintensity of 6.0-12.0 mW/cm² for 30 minutes to initiate the free-radicalpolymerization. The ensuing particles, typically of 800 nm to 20 micronsize were washed repeatedly in deionized water to eliminate anyunreacted monomers and to extract all the glucose. This process wascompleted in 4 days at 25-30 C.

The Wurster Bed Coating Process. In a suggested, typical coating studywith a Wurster bed, 10 g of the core material was charged into the bed.A 5% w/w dispersion of the CBIP particles prepared before in the amountof about 40 g (2 g of CBIP solids) at a feed rate of 2.4 min/min can besprayed on the core particle to form the 6 μm thick hydrogel layer.Then, 1 g of insulin was sprayed on the system at a feed rate of 2.4min/min. This process created a 2 μm thickness coating.

The factors governing the flow rate of the dispersion were the solidcontent of the dispersion and the stickiness of the material. The lowerthe solid content, the lower the feed rate to the nozzle, because ittook more time to dry the system. The dispersions can be fed to thespray nozzle by a peristaltic pump and atomized with a dried compressedair. The dispersion was continuously stirred during the coating process.

During all these processes, heated air (40° C.) was fed into the chamberat a flow rate of 0.25-0.75 m³/min to fluidize the particles. Thethickness of each layer was predetermined before the coating process.The calculation was based on the average size of the core and thethickness of the CBIP nanoparticles and insulin layers. The volume ofeach layer was calculated and adjusted by assuming 23% void fraction forspherical systems. The dry weight of the dispersion required to form thelayers for a particles was calculated from the known densities of thedispersion. The number of the particles could be calculated by dividingthe total weight of the core charged, which was 10 g by the weight of asingle core. Based on these calculations, the total dry weight of eachdispersion could be calculated.

Once the coating process was finished, the multilayered mimeticstructure was collected and weighed to determine the yield. Themultilayered mimetic structure was then sieved and the particle sizedistribution was analyzed. The multilayered mimetic structure was thendried under vacuum for 24 hr.

The feed rate of the latex dispersion depended on the properties of thelatex itself. Tacky dispersions, with low T_(g), required a lower feedrate. Dispersions with low solid content also required a low feed rateto allow the microcapsule to dry. The flow rate of the dried air used tofluidize the microcapsule controls the inertia of the microcapsule. Highinertia could prevent the microcapsule from agglomerating. However, highinertia could also break fragile microcapsule.

The temperature of the dried air at the inlet and outlet governs thetemperature of the bed. These temperatures control the rate of thedrying of the microcapsule. High temperature allows the microcapsule todry faster, and hence, allow a faster process. However, high temperaturecannot be applied for a dispersion system with low T_(g). Based on mypast experience on the Wurster coating process with other materials, theoperating conditions for the preparation of multilayer microcapsule inthis work must be around (i) feed rate of 2.4-3.0 min/min for MIPnanoparticle dispersion; (ii) feed rate of 2.4-4.0 min/min for insulindispersion; (iii) air flow rate of 0.25-0.75 m³/min; and (iv) air inlettemperature of 40° C.

EXAMPLE 2

A second prototype of the present invention was constructed for glucoserecognition followed by insulin release. The system was formed by usinga core, followed by a coating of the CBIP polymer in multiple layers.The core used was calcium carbonate crystals (size range 63-75 μm). TheMIP or recognitive layer consisted of a CBIP, hydroxy propyl cellulose(HPC) as a spacer and mannitol as a binder.

A CBIP coating was prepared by reacting 14 ml of acrylic acid (AA), 1.3g of IRGACURE® 184, 1-hydroxycyclohexyl phenyl ketone as an initiator,18 ml of dimethylsulfoxide (DMSO) as a solvent, and 7 g of poly(ethyleneglycol dimethacrylate) (PEGDMA, of PEG molecular weight of 200, 400 or1000) as a crosslinking agent. The system was reacted in the presence of10 g D-glucose.

The reaction was carried out in a 100 ml glass reactor placed under a UVsource (Dymax Ultraviolet Flood Cure System) and exposed to UV lightwith an intensity of 8.0-14.0 mW/cm² for 30 minutes to initiate thefree-radical polymerization. The ensuing particles, typically of 200 nmto 10 micron size were washed repeatedly in deionized water to eliminateany unreacted monomers and to extract all the glucose. This process wascompleted in 4 days at 25-30 C.

In a typical coating study with a Wurster bed, 10 g of the core materialwas charged into the bed. A 5% w/w dispersion of the CBIP partclesprepared before in the amount of about 40 g (2 g of CBIP solids) at afeed rate of 2.4 min/min can be sprayed on the core particle to form the6 μm thick hydrogel layer. Then, 1 g of insulin was sprayed on thesystem at a feed rate of 2.4 min/min. This process created a 2 μmthickness coating.

EXAMPLE 3 Formulation No. 2: Recognitive and Release-Triggering Systemswith Multiple Recognitive Coatings for Drug Delivery

The system in Example 2 can be developed into a multilayered ormulticoated device where each layer contains a recognitive hydrogelsprepared and applied to a core of drug by a spray coating technique(e.g., a Glatt GPCG, fluid bed coater, Glatt Air Techniques, Inc.,Ramsey, N.J.). In between each coating there are annular pouchescontaining finely encapsulated drug particles. Each coating (layer)expands because of the osmotic effect described before. Each hydrogelcoating layer is designed to rupture at preset times, thus releasing thesubsequent layer of encapsulated drug particles.

Using the above technology an active agent can be released in: multipledoses of the same drug at exactly the same time intervals; multipledoses of the same drug at varying intervals (e.g., first at 10 minutes,second at 15 minutes, third at 30 minutes, fourth at 5 minutes, etc);multiple doses of two different drugs at the same or varying timeintervals. These drugs maybe smaller molecular weight actives, peptides,proteins and others; multiple doses of several drugs or peptides orproteins at equal or varying intervals and/or release of an activefollowed by a second layer containing a sweetener to cover (mask) thebitter after taste of the first drug

EXAMPLE 4 Recognitive and Release-Triggering Laminates with MultipleRecognitive Layers for Drug or Other Active Delivery

A system can be developed based on the same principles as in FormulationNo. 2, but in the form of thin films (laminates) stacked on top of eachother. Thus, a multilayered device can be prepared where each filmcontains a recognitive hydrogel is prepared and applied to a core ofdrug by casting technique. In between each coating there are gapscontaining finely encapsulated drug particles. Each laminate expandsbecause of the osmotic effect described before. Each hydrogel laminate(film) is designed to rupture at preset times, thus releasing thesubsequent layer of encapsulated drug particles.

Using the above technology an active agent can be released in: multipledoses of the same drug at exactly the same time intervals; multipledoses of the same drug at varying intervals (e.g., first at 10 minutes,second at 15 minutes, third at 30 minutes, fourth at 5 minutes, etc);multiple doses of two different drugs at the same or varying timeintervals. These drugs maybe smaller molecular weight actives, peptides,proteins and others; multiple doses of several drugs or peptides orproteins at equal or varying intervals and/or release of an activefollowed by a second film containing a sweetener to cover (mask) thebitter after taste of the first drug.

EXAMPLE 5 Externally-Triggered Multilayered Skin-adherent Films forDelivery of Essential Oils or Cosmetic Actives or Bactericides orTopical Treatment Over-the-Counter Agents

A system can be developed based on the same principles as in Examples 2and 3, but in the form of thin films (laminates) stacked on top of eachother. In this case, however, the multilayered device can contain also afinal thin layer of a skin adhesive polymer (e.g., pressure sensitiveadhesive or poly(acrylic acid) or similar that can be used to attach thewhole system to the skin. The remaining part of the device is preparedwith each film containing a recognitive hydrogel prepared and applied toa core of active by a casting technique. In between each coating thereare gaps containing finely encapsulated particles of essential oils,cosmetic actives, bactericides or over-the-counter agents that can beused in local skin treatment. Each laminate expands because of theosmotic effect described before. Each hydrogel laminate (film) isdesigned to rupture at preset times, thus releasing the subsequent layerof encapsulated agent particles.

Using the above technology an active agent can release: multiple dosesof the same agent at exactly the same time intervals; multiple doses ofthe same agent at varying intervals (e.g., first at 10 minutes, secondat 15 minutes, third at 30 minutes, fourth at 5 minutes, etc); multipledoses of two different agents (e.g., a perfume compound and abactericide) at the same or varying time intervals; multiple doses ofseveral drugs agents at equal or varying intervals.

EXAMPLE 6 Sweat-Triggered Multilayered Skin-Adherent Films for Deliveryof Essential Oils or Cosmetic Actives

A system developed on principles similar to those of Examples 2-4, butin the form of multiple thin films (laminates) triggered by activesproduced by the human sweating process and releasing actives. Suchtriggering molecules will be epinephrine and related compounds. Themultilayered device will also contain a final thin layer of a skinadhesive polymer (e.g., pressure sensitive adhesive or poly(acrylicacid) or similar) that can be used to attach the whole system to theskin.

The remaining part of the device is prepared with each film containingan epinephrine-imprinted, recognitive hydrogel prepared and applied to acore of active by a casting technique. In between each coating there aregaps containing finely encapsulated particles of essential oils,cosmetic actives, bactericides or over-the-counter agents that can beused in local skin treatment. Each laminate expands because of theosmotic effect described before. Each hydrogel laminate (film) isdesigned to rupture at preset times, thus releasing the subsequent layerof encapsulated agent particles.

Using the above technology we can release: multiple doses of the sameagent at exactly the same time intervals; multiple doses of the sameagent at varying intervals (e.g., first at 10 minutes, second at 15minutes, third at 30 minutes, fourth at 5 minutes, etc); multiple dosesof two different agents (e.g., a perfume compound and a bactericide) atthe same or varying time intervals; and multiple doses of several drugsagents at equal or varying intervals.

EXAMPLE 7 Sweat-Triggered Multilayered Nanoparticles for Delivery ofEssential Oils or Cosmetic Actives

A system developed on principles similar to those of Examples 2-4, butin the form of multiple nanoparticles in the form of devices triggeredby actives produced by the human sweating process and releasing actives.Such triggering molecules will be epinephrine and related compounds.Such systems can be applied as a dry powder and can release at varioustimes, as late as 8 hours after application. They may contain expensiveperfume, essential oils, bactericides, etc.

EXAMPLE 8 Sweat-Triggered Multilayered Skin-Adherent Films for Deliveryof Oils, Lipids or Creams

Delivery of highly hydrophobic active agents using the MS/UT technology(after initial recognition) is extremely difficult as the lipidicstructures are extremely difficult to penetrate through hydrophilicgels. However, with the present technology a multivariate system can beprepared, where the encapsulated nanoparticles are wholly covered byhydrophobic coatings and reside in the space between consecutive layersof epinephrine-recognitive hydrogel films. A system was in the form ofmultiple thin films (laminates) triggered by actives produced by thehuman sweating process and releasing actives. Such triggering moleculeswill be epinephrine and related compounds. The multilayered device mayalso contain a final thin layer of a skin adhesive polymer (e.g.,pressure sensitive adhesive or poly(acrylic acid) or similar) that canbe used to attach the whole system to the skin.

The remaining part of the device is prepared with each film containingan epinephrine-imprinted, recognitive hydrogel prepared and applied to acore of active by a casting technique. In between each coating there aregaps containing finely encapsulated particles of creams that can be usedin local skin treatment. Each laminate expands because of the osmoticeffect described before. Each hydrogel laminate (film) is designed torupture at preset times, thus releasing the subsequent layer ofencapsulated agent particles.

The remaining part of the device is prepared with each film containingan epinephrine-imprinted, recognitive hydrogel prepared and applied to acore of active by a casting technique. In between each coating there aregaps containing finely encapsulated particles of creams that can be usedin local skin treatment. Each laminate expands because of the osmoticeffect described before. Each hydrogel laminate (film) is designed torupture at preset times, thus releasing the subsequent layer ofencapsulated agent particles.

EXAMPLE 9 Preparation of Recognitive Systems

Preparation of Films from Recognitive Polymers. The configurationalbiomimetic imprinted polymer (CBIP) recognitive films prepared in thiswork included a crosslinked copolymer of methacrylic acid (MAA 99%,inhibited with 100-250 ppm HQ, Sigma-Aldrich, St. Louis, Mo.) andethylene glycol dimethacrylate (EGDMA, stabilized, 98%, Acros Organics,N.J.). The reaction mixture was imprinted with D-glucose (A.C.S.reagent, Aldrich Chemical Co., Milwaukee, Wis.). The monomers werephotopolymerized in the presence of UV light with2,2-dimethoxy-2-phenylacetophenone (DMPA, 99%, Acros Organics) used asthe free-radical initiator. All solvents were of analytical grade.

MAA (10% w/w) and D-glucose (5% w/w) were added to a solution comprisedof a 3:1 ratio of ethanol to water (by volume) and were sonicated for 20minutes to evenly disperse the components. To this mixture, EGDMA (84%w/w) and DMPA (1% w/w) were added and mixed by mechanical shaking. Thismixture was then purged with N₂ in an oxygen-free environment (e.g., asealed glove box) for 30 minutes. After purging, the mixture was loadedinto a polymerization chamber (made from two 75×50×1 mm microscopeslides with a 200μ Teflon spacer placed in between, bound with binderclips around the edge) and polymerized under UV light (at 15 mW/cm2) for15-20 minutes. The resulting films were then removed from the glove boxand washed with Milli-Q DI-water for 24-48 hours (with the water beingchanged every 24 hours), after which they were removed and dried in adrying oven (with desiccant) for 24 hours. The resulting films were thenstored at room temperature for observation.

Preparation of Control Samples. The control samples (non-imprintedpolymer films) were prepared using precisely the same protocol as above,except for the absence of D-glucose from the mixture (i.e., no glucosewas added).

Configurational Biomimetic Imprinting in the Presence of Glucose. Thebiomedical applications for the novel systems are far reaching due to aunique development in the field known as molecular imprinting, a conceptupon which our CBIP systems are based. Molecular recognition viaimprinting is accomplished by having a certain template molecule (in ourcase, glucose) dispersed between the various monomers during the processof polymerization (FIG. 1). The monomers proceed to polymerize as usual,while the specific template is imprinted directly within the polymernetwork (FIG. 2). The polymer films are then washed, thus removing thetemplate molecule (e.g., glucose) and leaving behind a chemicallystereospecific site where the template molecule was once part of thenetwork (FIG. 3). When the film is subsequently dried, nanovacuoles arepresent, while the polymer's physical characteristics change (notably adecrease in overall size). When glucose is reintroduced to the system,the attachment of the molecule to the stereospecific site causes amechanical stress in the local region that will eventually lead torupturing; it is this template-specific recognition/rupture/deliverythat we seek to exploit in the form of novel drug delivery systems. Thetemplates used in this type of recognition can be extended to almost anymolecule; further investigation will reveal new applications for thistechnique.

Sample Preparation. After numerous different procedures or “recipes”were performed, a general procedure was selected. There were manyvariations to the results of each recipe and method. At present time,not all factors have been tested. Samples prepared with low amounts ofEGDMA as a crosslinker showed significant differences from those filmsmade with at least 25 wt % crosslinker (the preferred ones). Thecrosslinker must be present in a high enough amount to hold thecopolymer together. This was noticed from the films which remained fluidor tacky and glutinous after the polymerization time ended. Differentprocedures were also followed with different amounts of ethanol andwater. Adding more ethanol to the recipe seemed to help the filmspolymerize more but also sometimes made the films turn out white.

It must be noted though, that it is necessary to do also studies atlower crosslinking ratios because of the ability to imprint largerproteins in these systems. It is particularly necessary to appreciatethat such systems will be important for consumer applications whereadded lipids will be necessary.

The polymerization time (under the UV lamp in the glove box) made adifference as to how rigid and continuous each film was. The firstsamples were prepared by polymerization for approximately 15 to 20minutes according to the protocol described in Example 2. But many ofthe recognitive and non-recognitive films polymerized under theseconditions were still “sticky”. Some films were polymerized for longeramounts of time (up to 60 minutes).

There were other laboratory procedures that were followed beforecomplete polymerization of the films. First, the monomer solution wassonicated for at least 20 minutes before adding EGDMA and DMPA to ensurea mixture. Once the EGDMA and DMPA were added and sonicated again for afew seconds.

Characterization. Characterization of the prepared films was done byswelling studies, optical and electron microscopy, FTIR analysis anddifferential scanning calorimetry. We will report here only what hasbeen completed already. Microparticles and continuous films wereobserved under polarized light, normal light, and with fluoresceinisothiocyanate—(FITC)—glucose under fluorescent light. The swelling ofthe films was studied in solutions of different glucose concentrationsand DI water.

Swelling Studies. The purpose of these studies was to examine therecognitive and swelling characteristics of all recognitive samples.This was done because of numerous questions received by MimeticSolutions as to the ability of the recognitive samples to recognizetheir templates fast and rupture as a result of the stresses created.

For each swelling study, the recognitive continuous films were cut intodisks or squares between 0.13 mm and 0.22 mm thick. The disks were cutwith a cork-borer with a radius of approximately 8 mm; the squares werecut with a razor blade to approximately 9×9 mm². Before a swellingstudy, the weight, thickness, and dimensions of each sample wererecorded. The disks or squares were then immersed into beakers witheither DI water, 100 mg/dL glucose solution, 150 mg/dL glucose solution,or 200 mg/dL glucose solution. The DI water was used as a control to seeif the disks and squares would swell without glucose present. It wasfound that the samples exposed to DI water should not swell as much asthe samples placed in a glucose solution.

Every ten minutes, samples were removed by tweezers, blotted gently witha wipe, and weighed. The total time that each sample was out of solutionwas roughly one minute. All studies were done at 37 C. In addition, thepH of each solution was measured.

In all studies, the amount of penetrant uptake was calculated bysubtracting the dry weight of sample (polymer) from a later weight andthen dividing by the dry weight of sample. These values are a clearindication of a fast uptake by the recognitive system. These values wereplotted as a function of time and as a function of square root of time(see FIGS. 4-7). These Figures show that glucose is recognized by therecognitive samples, leading to fast binding. A very fast recognitionprocess is observed with glucose (100 mg/dl solution) with subsequentsaturation of the binding sites and reduction of the water uptake untila constant value is obtained, almost the same as for pure waterabsorption. Scientifically, this phenomenon can be interpreted in asimilar way as the action of solid catalysts. Once the active sites areoccupied, no further reaction can take place.

Numerous studies of recognition at higher glucose levels (e.g., 150mg/dL and 200 mg/dL) indicated that these samples fractured within a fewminutes (typically 13-20 minutes for thin films) after exposure to thesolutions. Indeed, graphs of penetrant uptake (1) versus time cannot bepresented as the weight decreased after being the samples were placed insolution because of the rupture or cracking.

FIG. 4 is a graph that shows the penetrant uptake of recognitive polymercontinuous films over time. The data points are the amount of penetrantuptake in Milli-Q deionized water (DI water) versus 100 mg/dL D-glucosein DI water. The films were cut into disks 8 mm in diameter and 0.12 mmthick; the initial weights were approximately 6 mg each. Measurementswere taken every 10 minutes.

FIG. 5 is a graph that shows the penetrant uptake of a recognitivepolymer continuous film versus the square root of time. The data pointsare amount of penetrant uptake in Milli-Q deionized water (DI water)versus 100 mg/dL D-glucose in DI water. The films were cut into disks 8mm in diameter and 0.12 mm thick; the weights were approximately 6 mgeach. Measurements were taken every 10 minutes.

FIG. 6 is a graph that shows the recognitive ratio of configurationalbiomimetic imprinted polymers (CBIP) in the presence of 100 mg per dLdeionized water compared to continuous films in the presence ofdeionized water. The films were cut into squares approximately 9 mm by 9mm and 0.22 mm thick. The penetrant uptake amount was obtained frommeasurements of mass every 10 minutes once the squares were placed inthe solutions of either deionized Water or glucose solution with 100 mgD-glucose per dL deionized water. The ratio is amount of penetrantuptake in the glucose solution to amount of penetrant uptake indeionized water.

Observation of Swelling and Recognitive Processes. A large number of therecognitive films were crushed using mortar and pestle or cut into 8 mmdisks using a cork borer in order to observe their response to glucosesolutions under the microscope. The swelling behavior of different CBIPand NIP (control) films was observed in DI water, and solutions of 100mg/dL glucose, 150 mg/dL glucose, 200 mg/dL glucose, 300 mg/dL ,FITC-glucose, and trypan blue. The samples were tested under normallight, polarized light, and fluorescent light (when using FITC-glucose).The particles (approximately 100-300 micrometers) were placed on amicroscope slide and pictures were taken before any solution was added.Then using either a spatula or a pipette, a drop or two of solution wereplaced on the samples. Effort was made to avoid capillarity effects.Still photographs were taken in succession to form videos. The purposeof using a trypan blue solution was to observe the liquid front movementinto the particles or films.

Observations Without Polarization. Swelling of particles and films wasdifficult to observe without polarized light. In general, the particlestended to agglomerate together when in solution, which could sometimesbe mistaken for swelling.

FIG. 8 shows typical recognitive response and swelling behavior ofparticles within a few seconds from exposure to the glucose-containingsolution. After one minute, the particles have started recognizingglucose, which creates internal stresses. The third panel of FIG. 8clearly shows the swollen particles that have formed from the largerparticles due to rupture.

FIG. 9 shows a very large number of particles produced by the samerecognitive process but using films containing a porosigen (see below)to produce large pores within the recognitive polymer.

FIG. 10 show stress lines formed during the first few seconds afteraddition of a glucose solution to a recognitive film produced asdescribed above. Clearly, these observed stress lines indicate theeffect of glucose on the CBIP system. These lines cannot be observed insimilar films exposed just to water or in NIP films (neat films, notimprinted).

FIG. 11 shows a typical sequence of stills from a video of therecognition/swelling and rupture of a thin film of a glucose-recognitiveCBIP film exposed to glucose. Clearly, the film ruptures with continuousextended cracks. This is a fundamental difference over the rupture ofCBIP particles (FIG. 8) that occurs in the form on numerous irregularlyshaped particles.

Observations of Moving Fronts with Dyes. Trypan blue was used in therecognitive solution to better delineate the fronts moving into all theparticles and to help us observe the recognition due to the motion ofthe glucose solution and associated swelling of particles and continuousfilms. FIG. 12 shows one such process with a clear indication of theposition of glucose fronts (darker area) as they penetrate in themicroparticles. This is a rather simple technique to verify frontpositions although it is less useful to identify stress lines (the clearindication of glucose action on the samples). This was achieved with thetechniques described below.

Observation of Recognitive Processes Using Polarized Light.

Polymer stresses as a result of recognition processes, polymer crazing,cracking or dissolution involve glucose or solvent transport into thepolymer followed by actual rupture and perhaps dissolution of thelatter. Numerous study techniques have been reported to study polymerstresses and dissolution. The simplest method is a gravimetric techniquewhere polymer specimens are immersed in a solvent and removed afterfixed time intervals. These specimens are then dried to remove residualsolvent and the thickness of the remaining film is measured. Thus, thetemporal evolution of the thickness is also obtained. Though simple,this method is tedious and almost impossible to use here.

Laser interferometry has been used to measure the polymer dissolutionrate of thin films. The technique has found applicability especially infollowing the dissolution of microlithographic masking layers. In thisstudy, a recognitive polymer coated on a silicon wafer is placed in asolvent. When a laser beam impinges upon the polymer surface, it splitsinto two beams: one is reflected by the surface and the other penetratesthe thin polymer film and is reflected by the silicon wafer. The twobeams interfere with each other, providing a measure of the change ofthe polymer film thickness as a function of time. This technique is alsolimited to those polymers that give a negligible gel layer because goodinterference cannot be obtained if the gel layer thickness issignificant.

Differential refractometry can also be used to measure the polymerdissolution rate. In this study, a polymer sample is immersed in aglucose solution in a special container, equipped with a differentialrefractometer and an agitator. Polymer cracking and dissolution arefollowed by measuring the refractive index of the solution. Using thismethod, it is also possible to measure induction times, which are thetimes necessary for a build-up of a swollen surface layer. Thedifferential refractometry technique can be used even in the presence ofgel layers. However the thickness of the gel layer cannot be measuredsimultaneously.

To measure both the recognition and CBIP polymer dissolution rate andthe gel layer thickness as well as to investigate polymer morphologyduring dissolution, techniques using optical microscopy have been used.The apparatus consists of an optical microscope and a sample cellcontaining a CBIP polymer sample engulfed by a matrix inert to thesolvent and sandwiched between two glass slides. As the contrast betweenthe different layers was usually poor, dyes are resorted to in theglucose solution.

To improve the contrast between the different layers, we designed anoptical microscopy apparatus with modifications in the sample cell andthe optical design, which obviated the need for the dye tracer. Bychanging the angle of illumination of the sample, we found that thecontrast achieved between the different layers in the recognitive,cracking/rupturing or dissolving polymer was sufficient for theirresolution. The optimum angle of illumination depended on the refractiveindices of the glucose solution and the CBIP polymer, and also on thesample thickness. The sample cell design was also modified to improvethe flow rate control and to allow greater precision in measuring themotions of the boundaries of the different layers.

Thus, techniques using optical microscopy are good tools for observingthe different layers as well as the possible crazing or cracking at theinterface of a dissolving CBIP polymer. Under polarized light,glucose-sensitive stress lines and fractures appear in the presence of aglucose solution. When the polarizer and analyzer of the polarizedmicroscope are crossed, the only light that can be seen is frombirefringence (as shown in FIG. 13 from a single particle before it hasstarted rupturing). This birefringence is the result of stress linescaused by glucose expanding the polymer particles. A typical sampleshowing birefringence is seen in FIG. 14 that presents a CBIP particleas it has started to rupture and crack. In this particular study, theruptured particles appeared after 2 min and 55 sec.

Observation of Recognitive Processes Using Fluorescent Markers.FITC-glucose provided one of the best ways to observe the recognitiveprocess as well as the swelling of the particles and continuous films,because fluorescence can be readily detected. We have not completedstudies in fluorescent light, but FIGS. 15 and 16 show some of theresults. The brightness of the particles corresponds to the stresses andswelling caused by the FITC-glucose (FIG. 15). The swelling generallyoccurred within 3-5 minutes after contact with the glucose solution.Recognition and stress development was very much dependant on particlesize. Particle of 10-15 microns were recognitive within 10 seconds!

Particles appeared to burst in the presence of FITC-glucose (FIG. 16).These bursts could probably be FITC-glucose solution “bubbles” burstingwithin the particles after they have had enough stress from surroundingsolution. This proves that the recognition of glucose does in fact leadto enough mechanical stress to cause the recognitive particles to breakapart. This is exactly what the ultimate goal of these studies is: tohave a recognitive layer with a glucose template that will burst uponrecognition of an abnormal level of glucose and release insulin.

FIG. 17 shows, at a high level, the basic combinations of the presentinvention. As the skilled artisan will appreciate, the molecule that isused to form the micro or nanovacuoles (shown here as the analyte) canbe any of a wide variety of molecules (or combinations thereof) that canbe recognized by the polymeric network. Upon exposure the analyte (i.e.,the recognition event), a polymeric transduction event occurs in whichone or more of the listed forces (or even additional forces) trigger adissociation, degradation, decomposition or otherwise reduce thestructural integrity of the polymeric network, thereby triggering thedelivery of a payload. One example of a payload may even be a lower orsubsequent layer of polymeric network, which may also include a core onwhich the layers may be disposed.

FIG. 18 is a diagram that shows mixing multilayered mimetic structureswith different release profiles that allows the system to be tailored tofit any release profiles. Mixing four different microcapsules allowedthe system to rupture every 3 h for 4 days instead of a system thatruptured every 12 h for 4 days or a system that ruptured every 3 h forone day.

EXAMPLE 10 Limited Swelling of the Recognitive Polymeric Network inSolvent Alone

The configurational biomimetic imprinted polymer (CBIP) recognitivefilms prepared in this work included a crosslinked copolymer of 4.0 mlwater, 4.5 ml ethanol, 60 mg glucose, 0.42 g MAA, 3.1 g TEGDMA,optionally a few drops of ethanol. Next, 50 mg DMPA was added. Thesolution is degassed for 4 minutes and loaded. The polymer is formed byUV irradiation for 5 minutes. The polymer opaque while film is washedwith water and removed from the slide. Washing is as describedhereinabove.

FIG. 19 shows glucose/water uptake in which glucose solution caused thesamples to fall apart quite quickly. The graph shows that the polymerswelled between 5-15 percent (±3%) in the presence of a solvent (water)alone. Upon exposure to the solvent and the analyte the polymericnetwork burst to release the payload. At lower glucose concentrations itis expected that the release can run the time course to equilibrium.

It is contemplated that any embodiment discussed in this specificationcan be implemented with respect to any method, kit, reagent, orcomposition of the invention, and vice versa. Furthermore, compositionsof the invention can be used to achieve methods of the invention.

It will be understood that particular embodiments described herein areshown by way of illustration and not as limitations of the invention.The principal features of this invention can be employed in variousembodiments without departing from the scope of the invention. Thoseskilled in the art will recognize, or be able to ascertain using no morethan routine experimentation, numerous equivalents to the specificprocedures described herein. Such equivalents are considered to bewithin the scope of this invention and are covered by the claims.

All publications and patent applications mentioned in the specificationare indicative of the level of skill of those skilled in the art towhich this invention pertains. All publications and patent applicationsare herein incorporated by reference to the same extent as if eachindividual publication or patent application was specifically andindividually indicated to be incorporated by reference.

The use of the word “a” or “an” when used in conjunction with the term“comprising” in the claims and/or the specification may mean “one,” butit is also consistent with the meaning of “one or more,” “at least one,”and “one or more than one.” The use of the term “or” in the claims isused to mean “and/or” unless explicitly indicated to refer toalternatives only or the alternatives are mutually exclusive, althoughthe disclosure supports a definition that refers to only alternativesand “and/or.” Throughout this application, the term “about” is used toindicate that a value includes the inherent variation of error for thedevice, the method being employed to determine the value, or thevariation that exists among the study subjects.

As used in this specification and claim(s), the words “comprising” (andany form of comprising, such as “comprise” and “comprises”), “having”(and any form of having, such as “have” and “has”), “including” (and anyform of including, such as “includes” and “include”) or “containing”(and any form of containing, such as “contains” and “contain”) areinclusive or open-ended and do not exclude additional, unrecitedelements or method steps.

The term “or combinations thereof” as used herein refers to allpermutations and combinations of the listed items preceding the term.For example, “A, B, C, or combinations thereof” is intended to includeat least one of: A, B, C, AB, AC, BC, or ABC, and if order is importantin a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC, or CAB.Continuing with this example, expressly included are combinations thatcontain repeats of one or more item or term, such as BB, AAA, MB, BBC,AAABCCCC, CBBAAA, CABABB, and so forth. The skilled artisan willunderstand that typically there is no limit on the number of items orterms in any combination, unless otherwise apparent from the context.

All of the compositions and/or methods disclosed and claimed herein canbe made and executed without undue studyation in light of the presentdisclosure. While the compositions and methods of this invention havebeen described in terms of preferred embodiments, it will be apparent tothose of skill in the art that variations may be applied to thecompositions and/or methods and in the steps or in the sequence of stepsof the method described herein without departing from the concept,spirit and scope of the invention. All such similar substitutes andmodifications apparent to those skilled in the art are deemed to bewithin the spirit, scope and concept of the invention as defined by theappended claims.

REFERENCES

1. E. S. Golub and D. R. Green, Immunology: A Synthesis. 1991,Sunderland, M A: Sinauer Assoc.

2. F. Breinl and F. Haurowitz, Chemical Investigation of the Precipitatefrom Hemoglobin and Anti-hemoglobin Serum and Remarks on the Nature ofAntibodies. Z. Physiol. Chem., 1930. 192: p. 45.

3. L. Pauling, A Theory of the Structure and Process of Formation ofAntibodies. J. Am. Chem. Soc., 1940. 62: p. 2643-2657.

4. N. K. Jeme, The Natural Selection Theory of Antibody Formation. Proc.Natl. Acad. Sci. USA, 1955. 41: p. 849.

D. W. Talmage, Allergy and Immunology. Ann. Rev. Med., 1957. 8: p. 239.

6. F. M. Burnet, A Modification of Jeme's Theory of Antibody ProductionUsing the Concept of Clonal Selection. Aust. J. Sci., 1957. 20: p. 67.

7. D. R. Davies and S. Chacko, Antibody Structure. Accounts Chem. Res.,1993. 26: p. 421-427.

8. N. K. Jerne, The Generative Grammar of the Immune System, in NobelLectures, Physiology or Medicine 1981-1990, J. Lindsten, Editor. 1993,World Scientific Publishing Co.: Singapore. p. 211-225.

9. G. Köhler and C. Milstein, Continuous cultures of fused cellssecreting antibody of predefined specificity. Nature, 1975. 256: p.495-497.

10. G. J. F. Köhler, Derivation and Diversification of MonoclonalAntibodies, in Nobel Lectures, Physiology or Medicine 1981-1990, J.Lindsten, Editor. 1993, World Scientific Publishing Co.: Singapore. p.228-243.

11. C. Milstein, From the Structure of Antibodies to the Diversificationof the Immune Response, in Nobel Lectures, Physiology or Medicine1981-1990, J. Lindsten, Editor. 1993, World Scientific Publishing Co.:Singapore. p. 248-270.

12. R. B. Merrifield, Solid Phase Peptide Synthesis. I. The Synthesis ofa Tetrapeptide. J. Am. Chem. Soc., 1963. 85: p. 2149-2154.

13. R. B. Merrifield, Solid Phase Peptide Synthesis. II. The Synthesisof Bradykinin. J. Am. Chem. Soc., 1964. 86: p. 304.

14. R. B. Merrifield, Solid-Phase Peptide Synthesis, III. An ImprovedSynthesis of Bradykinin. Biochem., 1964. 3: p. 1385-1390.

15. B. Merrifield, Solid Phase Synthesis, in Nobel Lectures, Chemistry1981-1990, T. Frängsmyr, Editor. 1992, World Scientific Publishing Co.:Singapore. p. 149-175.

16. K. Kirshenbaum, A. E. Barron, R. A. Goldsmith, P. Armand, E. K.Bradley, K. T. V. Truong, K. A. Dill, F. E. Cohen, and R. N. Zuckermann,Sequence-specific polypeptoids: A diverse family of heteropolymers withstable secondary structure. Proc. Natl. Acad. Sci. USA, 1998. 95: p.4303-4308.

17. M. Egholm, E. Buchardt, P. E. Nielsen, and R. H. Berg, Peptidenucleic acids (PNA). Oligonucleotide analogues with an achiral peptidebackbone. J. Am. Chem. Soc., 1992. 114: p. 1895-1897.

18. G. P. Dado and S. H. Gellman, Intramolecular Hydrogen Bonding inDerivatives of Beta-Alanine and Gamma-Amino Butyric Acid: Model Studiesfor the Folding of Unnatural Polypeptide Backbones. J. Am. Chem. Soc.,1994. 116: p. 1054-1062.

19. D. H. Appella, L. A. Christianson, I. L. Karle, D. R. Powell, and S.H. Gellman, Peptide Foldamers: Robust Helix Formation in a New Family ofBeta-Amino Acid Oligomers. J. Am. Chem. Soc., 1996. 118: p. 13071-13072.

20. K. Yue and K. A. Dill, Inverse Protein Folding Problem: DesigningPolymer Sequences. Proc. Natl. Acad. Sci. USA, 1992. 89: p. 4163-4167.

21. S. H. Gellman, Foldamers: A Manifesto. Accounts Chem. Res., 1998.31: p. 173-180.

22. K. Kirshenbaum, R. N. Zuckermann, and K. A. Dill, Designing polymersthat mimic biomolecules. Current Opinion in Structural Biology, 1999. 9:p. 530-535.

23. P. Koehl and M. Levitt, De Novo Protein Design. I. In Search ofStability and Specificity. J. Mol. Biol., 1999. 293: p. 1161-1181.

24. P. Koehl and M. Levitt, De Novo Protein Design. II. Plasticity inSequence Space. J. Mol. Biol., 1999. 293: p. 1183-1193.

25. E. I. Shakhnovich and A. M. Gutin, Engineering of stable andfast-folding sequences of model proteins. Proc. Natl. Acad. Sci. USA,1993. 90: p. 7195-7199.

26. V. S. Pande, A. Y. Grosberg, and T. Tanaka, Folding Thermodynamicsand kinetics of imprinted renaturable heteropolymers. J. Chem. Phys.,1994. 101(9): p. 8246-8257.

27. V. S. Pande, A. Y. Grosberg, and T. Tanaka, Thermodynamic procedureto synthesize heteropolymers that can renature to recognize a giventarget molecule. Proc. Natl. Acad. Sci. USA, 1994. 91: p. 12976-12979.

28. V. S. Pande, A. Y. Grosberg, and T. Tanaka, Phase diagram ofheteropolymers with an imprinted conformation. Macromolecules, 1995. 28:p. 2218-2227.

29. V. S. Pande, A. Y. Grosberg, and T. Tanaka, How to Create Polymerswith Protein-Like Capabilities: A Theoretical Suggestion. Physica D,1997. 107: p. 316-321.

30. K. Mosbach and O. Ramstrom, The Emerging Technique of MolecularImprinting and its Future Impact on Biotechnology. Biotechnol., 1996.14: p. 163-170.

31. P. A. G. Cormack and K. Mosbach, Molecular imprinting: recentdevelopments and the road ahead. Reactive and Functional Polymers, 1999.41: p. 115-124.

32. K. Mosbach, Toward the next generation of molecular imprinting withemphasis on the formation, by direct molding, of compounds withbiological activity(biomimetics). Anal. Chim. Acta, 2001. 435: p. 3-8.

33. M. Komiyama, T. Takeuchi, T. Mukawa, and H. Asanuma, MolecularImprinting From Fundamentals to Applications. 2003, Weinheim, Germany:Wiley-VCH.

34. G. Wulff, A. Sarhan, and K. Zabrocki, Enzyme-Analogue Built Polymersand Their Use for the Resolution of Racemates. Tetrahedron Letters,1973. 44: p. 4329-4332.

35. C. Yu and K. Mosbach, Influence of mobile phase composition andcross-linking density on the enantiomeric recognition properties ofconfigurationally biomimetic imprinted polymers. J. Chromatogr. A, 2000.888: p. 63-72.

36. H. S. Andersson, J. G. Karlsson, S. A. Piletsky, A.-C. Koch-Schmidt,K. Mosbach, and I. A. Nicholls, Study of the nature of recognition inconfigurationally biomimetic imprinted polymers, II. Influence ofmonomer-template ratio and sample load on retention and selectivity. J.Chromatogr. A, 1999. 848: p. 39-49.

37. D. Kriz, C. B. Kriz, L. I. Anderson, and K. Mosbach, Thin-LayerChromatography Based on the Molecular Imprinting Technique. Anal. Chem.,1994. 66: p. 2636-2639.

38. J. Cederfur, Y. Pei, M. Zihui, and M. Kempe, Synthesis and Screeningof a Configurationally biomimetic imprinted Polymer Library Targeted forPenicillin G. J. Comb. Chem., 2003. 5: p. 67-72.

39. M. Kempe and K. Mosbach, Separation of amino acids, peptides andproteins on configurationally biomimetic imprinted stationary phases. J.Chromatogr. A, 1995. 691: p. 317-323.

40. K. Haupt, K. Noworytab, and W. Kutner, Imprinted polymer-basedenantioselective acoustic sensor using a quartz crystal microbalance.Anal. Commun., 1999. 36.

41. C. Liang, H. Peng, A. Zhou, L. Nie, and S. Yao, Molecular imprintingpolymer coated BAW bio-mimic sensor for direct determination ofepinephrine. Anal. Chim. Acta, 2000. 415: p. 135-141.

42. J. Z. Hilt, M. E. Byrne, and N. A. Peppas. Novel Biomimetic PolymerNetworks: Development and Application as Selective Recognition Elementsfor Biomolecules at the Micro-/Nanoscale. in AIChE Nanoscale Science andEngineering Topical Conference Proceedings. 2003. San Francisco, Calif.

43. D. K. Robinson and K. Mosbach, Molecular imprinting of a transitionstate analogue leads to a polymer exhibiting esterolytic activity. J.Chem. Soc. Chem. Commun., 1989. 14: p. 969 - 970.

44. B. Sellergren and K. J. Shea, Enantioselective ester hydrolysiscatalyzed by imprinted polymers. Tetrahedron-Asymmetry, 1994. 5: p.1403-1406.

45. R. N. Karmalkar, M. G. Kulkarni, and R. A. Mashelkar,Configurationally biomimetic imprinted Hydrogels ExhibitChymotrypsin-like Activity. Macromolecules, 1996. 29: p. 1366-1368.

46. O. Ramström and K. Mosbach, Synthesis and catalysis byconfigurationally biomimetic imprinted materials. Curr. Opin. Chem.Biol., 1999. 3: p. 759-764.

47. L. Andersson, B. Sellergren, and K. Mosbach, Imprinting of AminoAcid Derivatives in Macroporous Polymers. Tetrahedron Letters, 1984.25(45): p. 5211-5214.

48. G. Vlatakis, L. J. Andersson, R. Muller, and K. Mosbach, Drug assayusing antibody mimics made by molecular imprinting. Nature, 1993. 361:p. 645-647.

49. R. A. Bartsch and M. Maeda, eds. Molecular and Ionic Recognitionwith Imprinted Polymers. ACS Symposium Series. 1998, ACS: Washington,D.C.

50. L. I. Andersson, Molecular Imprinting as an Aid to Drug Bioanalysis,in Drug Development Assay Approaches, Including Molecular Imprinting andBiomarkers, E. Reid, H. M. Hill, and I. D. Wilson, Editors. 1998, TheRoyal Society of Chemistry: Cambridge, UK.

51. G. Odian, Principles of Polymerization. 1991, New York: Wiley.

52. G. Wulff, J. Vietmeier, and H.-G. Poll, Enzyme-analogue builtpolymers, 22: Influence of the nature of the crosslinking agent on theperformance of imprinted polymers in racemic resolution. Makromolekul.Chem., 1987. 188: p. 731-740.

53. B. D. Ratner, The Engineering of Biomaterials Exhibiting Recognitionand Specificity. J. Mol. Recognition, 1996. 9: p. 617-625.

54. N. A. Peppas and R. Langer, Advances in Biomaterials, Drug Delivery,and Bionanotechnology. AIChE J., 2003. 49(12): p. 2990-3006.

55. M. E. Byrne, K. Park, and N. A. Peppas, Molecular imprinting withinhydrogels. Adv. Drug Deliver. Rev., 2002. 54(1): p. 149-161.

56. M. E. Byrne, K. Park, and N. A. Peppas. Biomimetic Networks forSelective Recognition of Biomolecules. 2002: Materials Research Society.

57. J. Z. Hilt, A. K. Gupta, R. Bashir, and N. A. Peppas, UltrasensitiveBiomems Sensors Based on Microcantilevers Patterned with EnvironmentallyResponsive Hydrogels. Biomedical Microdevices, 2003. 5(3): p. 177-184.

58. E. Oral and N. A. Peppas, Responsive and recognitive hydrogels usingstar polymers. J. Biomed. Mater. Res. A, 2004. 68: p. 439-447.

59. P. Parmpi and P. Kofinas, Biomimetic glucose recognition usingconfigurationally biomimetic imprinted hydrogels. Biomaterials, 2004.25: p. 1969-1973.

60. L. D. V. Bolisay, J. F. March, W. E. Bentley, and P. Kofinas,Separation of baculoviruses using configurationally biomimetic imprintedpolymer hydrogels. Mat. Res. Soc. Symp. Proc., 2004. 787: p.G3.1/1-G3.1/5.

61. D. R. Shnek, D. W. Pack, D. Y. Sasaki, and F. H. Arnold, SpecificProtein Attachment to Artificial Membranes via Coordination toLipid-Bound Copper(I1). Langmuir, 1994. 10: p. 2382-2388.

62. M. Kempe, M. Glad, and K. Mosbach, An approach towards surfaceimprinting using the enzyme ribonuclease A. J. Mol. Recognition, 1995.8(1-2): p. 35-39.

63. L. I. Andersson, R. Muller, G. Vlatakis, and K. Mosbach, Mimics ofthe Binding Sites of Opiod Receptors Obtained by Molecular Imprinting ofEnkephalin and Morphine. Proc. Natl. Acad. Sci. USA, 1995. 92: p.4788-4792.

64. D. L. Venton and E. Gudipati, Influence of protein on polysiloxanepolymer formation: evidence for induction of complementaryprotein-polymer interactions. Biochim. Biophys. Acta, 1995. 1250: p.126-136.

65. A. Rachkov and N. Minoura, Towards Configurationally biomimeticimprinted Polymers Selective to Peptides and Proteins. The EpitopeApproach. Biochimica et Biophysica Acta, 2001. 1544: p. 255-266.

66. L. J. Harris, S. B. Larson, K. W. Hasel, and A. McPherson, RefinedStructure of an Intact IgG2a Monoclonal Antibody. Biochem., 1997. 36: p.1581-1597.

67. H. M. Berman, J. Westbrook, Z. Feng, G. Gilliland, T. N. Bhat, H.Weissig, I. N. Shindyalov, and P. E. Bourne, The Protein Data Bank.Nucleic Acids Res., 2000. 28: p. 235-242.

68. D. R. Burton, Monoclonal Antibodies from Combinatorial Libraries.Accounts Chem. Res., 1993. 26: p. 405-411.

69. J. Tormo, D. Blaas, N. R. Parry, D. Rowlands, D. Stuart, and I.Fita, Crystal Structure of a Human Rhinovirus Neutralizing AntibodyComplexed with a Peptide Derived from Viral Capsid Protein VP2. EMBO J.,1994. 13: p. 2247-2256.

70. L. Stryer, Biochemistry. 4th ed. 1995, New York: W. H. Freeman.

71. K. Mosbach, K. Haupt, X.-C. Liu, P. A. G. Cormack, and O. Ramstrom,Molecular Imprinting: Status Artis et Quo Vadere, in Molecular and IonicRecogniton with Imprinted Polymers, R. A. Bartsch and M. Maeda, Editors.1998, American Chemical Society: Washington, D.C.

ADDITIONAL REFERENCES

Burt S. Essential Oils: their antibacterial properties and potentialapplications in food—a review. International Journal of FoodMicrobiology 94 (2004) pp. 223-253

Canal, T.; Peppas, N. A. J. Biomed. Mater. Res. (1989) 23, 1183.

Chang C. P., Dobashi T. Preparation of alginate complex capsulescontaining eucalyptus essential oil and its controlled release. Colloidsand Surfaces B: Biointerfaces 32 (2003) pp.257-262

Duclairoir C., Orecchioni A. M., Depraetere P., Osterstock F., NakacheE. Evaluation of gliadins nanoparticles as drug delivery systems: astudy of three different drugs. International Journal of Pharmaceutics,253 (2003) p. 133-144

Franzios G., Mirotsou M., Hatziapostolou E., Kral J., Scouras Z. G.,Mavragani-Tsipidou P. Insecticidal and genotoxic activities of mintessential oils. Journal of Agricultural and Food Chemistry, 45 (1997)pp. 2690-2694

Hartmans K. J., Diepenhorst P. The use of carvone as a sprout inhibitorfor potatoes. Potato Research, 37 (1994) pp. 445-446

Hartmans K. J., Lenssen J. M., De Vries R. G. Use of talent (carvone) asa sprout growth regulator of seed potatoes and the effect on stm andtuber number, Potato Research, 41 (1998) pp. 190-191

Muller N. J. United States Patent Application 20020071877 (2002)

Peppas N. A., Am Ende D. J. Controlled release of perfumes frompolymers. II. Incorporation and release of essential oils from glassypolymers, Journal of Applied Polymer Science, Vol. 66 (1997) pp. 509-513

Peppas N. A., Brannon-Peppas L. Controlled Release of fragrance frompolymers I. Thermodynamic analysis. Journal of Controlled Release 40(1996) 245-250

Sanchez I. C. Equilibrium distribution of a minor constituent between apolymer and its environment, in Durability of Macromolecular Materials,R. K. Eby, ed. ACS Symp. Ser. Vol. 95, American Chemical Society,Washington, DC, 1979, pp. 171-181

Sanchez I. C., Chang S. S. and Smith L. E. Migration models for polymeradditives, Polym. News 6 (1980) 249-256

Secouard S., Malhiac C., Grisel M., Decroix B. Release of limonene frompolysaccharide matrices: viscosity and synergy effect. Food Chemistry 82(2003) 227-234

1. A composition comprising: a molecule-imprinted polymeric networkcomprising a polymer, a micromolecular systems, a hydrogel, a gel or acomposite comprising one or more micro- or nanovacuoles or pores,wherein the nanovacuoles recognize a specific molecule and subsequentcontact with the molecule creates internal stresses that rupture thepolymeric network at the micro-or nanovacuoles.
 2. The composition ofclaim 1, wherein rupture of the polymeric network is due to: osmosisupon recognition and binding of the molecule leading to rupture due toswelling; change of the solubility of the polymeric network leading topolymer dissolution; local temperature changes leading to expansion ofthe polymeric network and combinations thereof.
 3. The composition ofclaim 1, wherein the composition is loaded with one or more activeagents to form an active agent-loaded, molecule-imprinted network. 4.The composition of claim 3, wherein the molecule-imprinted network isimprinted in a carbohydrate polymer of glycosidic type mono-sugarrepeative units, galactomannans, pectins, alginates, carrageenans andxanthan gum that are linear or branched, neutral or anionic andcombinations thereof.
 5. The composition of claim 3, wherein the activeagent-loaded, molecule-imprinted network is adapted for pharmaceuticals,medical agents, food components, detergents, bleaches, fabric softeners,fragrances, cosmetic products, air fresheners, room deodorant devices,perfumed substrates, perfumed plastics, pet collars, food and cosmeticapplications, hydrocolloids extracted from plants, seaweeds or animalcollagen, produced by microbial synthesis, and comprise polysaccharides,proteins, for personal care selected from hair care (shampoos, hairmousses, styling agents); skin care (body lotions, vitamin e, aloevera); bath products (moisture-triggered release products); bodypowders; toilet soap (milled or poured, ionic strength-triggeredrelease), household products selected from laundry care; paper products;specialty cleaners (chlorinated cleaners, scouring pads, effervescenttoilet bowl cleaner powders); air fresheners, cosmetics and treatmentproducts selected from lipstick; eye-liner; foundation; base; blush;mascara; eye shadow; lip liner; facial powder; consealer; facial cream;make-up remover; mascara remover; make-up; skin treatments, a fragranceselected from cologne; perfume; sampling; antiperspirant; deodorant;anti-dandruff shampoos; athlete foot products and combinations thereof.6. The composition of claim 3, wherein the active agent-loaded,molecule-imprinted network is encapsulated.
 7. The composition of claim3, wherein the active agent-loaded, molecule-imprinted network isencapsulated and comprises a surfactant, a bleaching agent, a corrosioninhibitor, a sudsing modifier, a fluorescent whitening agent, one ormore enzymes, an anti redeposition agent, a color, one or more additivesand combinations thereof.
 8. The composition of claim 3, wherein theactive agent is released upon a change in solubility, pressure, a pHshift, a change in temperature, a temperature increase, enzymaticbreakdown, diffusion and combinations thereof.
 9. The composition ofclaim 1, wherein the polymeric recognitive network is formed into one ormore layers.
 10. The composition of claim 1, wherein the polymericrecognitive network is formed into one or more layers, each of whichrecognizes one or more different molecules and each of which provides abarrier to the release of one or more different active or inert agentsor both.
 11. The composition of claim 1, wherein the polymericrecognitive network is formed into a sphere, film, planar,semi-spherical, cylinder, rod, hemispheres, conical, hemi-cylinders andcombination thereof.
 12. The composition of claim 1, wherein thepolymeric recognitive network is at least partially porous.
 13. Anactive agent-loaded, molecule-imprinted polymeric network comprising:two or more active agent loaded, aggregated polymeric or gelnanoparticles or microparticles comprising micro- or nanovacuoles ormicro- or nanopores previously imprinted with a molecule, wherein one ormore pre-determined molecules bind specifically to the micro- ornanovacuoles or micro- or nanopores and contact with the moleculecreates internal stresses that rupture the network at the micro- ornanovacuoles or micro- or nanopores thereby releasing one or more activeagents loaded into the active agent-loaded, molecule-imprinted polymericnetwork.
 14. A method of making a recognitive release system comprising:selecting one or more molecules for recognition; forming micro- ornano-vacuoles in a polymeric recognitive network about the one or moremolecules; removing the molecule from the micro- or nano-vacuoles; andcoating one or more active agents with a polymeric recognitive network,wherein the one or more active agents release upon contact by thepolymeric recognitive network with its cognate molecule.
 15. The methodof claim 14, wherein the polymeric recognitive network is formed about acore.
 16. The method of claim 14, wherein the polymeric recognitivenetwork is coated into multiple layers.
 17. The method of claim 14,wherein the polymeric recognitive network is coated by a Wurster coatingmethod.
 18. The method of claim 14, wherein the polymeric recognitivenetwork is further encapsulated in a capsule, caplet, softgel, gelcap,suppository, film, granule, gum, insert, pastille, pellet, troche,lozenge, disk, poultice or wafer.
 19. The method of claim 14, whereinthe polymeric recognitive network is formed into one or more layers,each of which recognizes one or more different molecules and each ofwhich provides a barrier to the release of one or more different activeor inert agents or both.
 20. The method of claim 14, wherein thepolymeric recognitive network is formed into a sphere, film, planar,semi-spherical, cylinder, rod, hemispheres, conical, hemi-cylinders andcombination thereof.
 21. The method of claim 14, wherein the polymericrecognitive network is at least partially porous.
 22. A method of makinga polymeric recognitive network comprising: selecting one or moretargets for recognition; forming micro- or nano-vacuoles in thepolymeric recognitive network about the one or more targets; embeddingwithin the polymeric recognitive network one or more active agents forrelease upon dissociation of the polymeric recognitive network; andremoving the targets from the micro- or nano-vacuoles, whereinsubsequent binding of the target to the micro- or nano-vacuoles causesdisruption of the polymeric recognitive network and release of the oneor more active agents.
 23. A method of loading an active agent in amolecule-imprinted network comprising: mixing the active agent to themolecule-imprinted network at or about room temperature; heating themixture to about 40° C. to swell the molecule-imprinted network, whereinthe active agent diffuses into the molecule-imprinted network; andcooling the mixture to about room temperature, wherein themolecule-imprinted network collapses and traps the active agent withinthe molecule-imprinted network.
 24. The method of claim 23, wherein theactive agent-loaded, molecule-imprinted network is adapted forpharmaceuticals, medical agents, food components, detergents, bleaches,fabric softeners, fragrances, cosmetic products, air fresheners, roomdeodorant devices, perfumed substrates, perfumed plastics, pet collars,food and cosmetic applications, hydrocolloids extracted from plants,seaweeds or animal collagen, produced by microbial synthesis, andcomprise polysaccharides, proteins, for personal care selected from haircare (shampoos, hair mousses, styling agents); skin care (body lotions,vitamin e, aloe vera); bath products (moisture-triggered releaseproducts); body powders; toilet soap (milled or poured, ionicstrength-triggered release), household products selected from laundrycare; paper products; specialty cleaners (chlorinated cleaners, scouringpads, effervescent toilet bowl cleaner powders); air fresheners,cosmetics and treatment products selected from lipstick; eye-liner;foundation; base; blush; mascara; eye shadow; lip liner; facial powder;consealer; facial cream; make-up remover; mascara remover; make-up; skintreatments, a fragrance selected from cologne; perfume; sampling;antiperspirant; deodorant; anti-dandruff shampoos; athlete foot productsand combinations thereof.
 25. The method of claim 23, wherein themolecule-imprinted network comprises a carbohydrate polymer ofglycosidic type mono-sugar repeative units, galactomannans, pectins,alginates, carrageenans and xanthan gum that are linear or branched,neutral or anionic and combinations thereof.
 26. The method of claim 23,wherein the active agent-loaded, molecule-imprinted network isencapsulated.
 27. The method of claim 23, wherein the active agent isreleased upon a change in solubility, pressure, a pH shift, a change intemperature, a temperature increase, enzymatic breakdown, diffusion andcombinations thereof.
 28. The method of claim 23, wherein the polymericrecognitive network is formed into one or more layers.
 29. The method ofclaim 23, wherein the polymeric recognitive network is formed into oneor more layers, each of which recognizes one or more different moleculesand each of which provides a barrier to the release of one or moredifferent active or inert agents or both.
 30. The method of claim 23,wherein the polymeric recognitive network is formed into a sphere, film,planar, semi-spherical, cylinder, rod, hemispheres, conical,hemi-cylinders and combination thereof.
 31. The method of claim 23,wherein the polymeric recognitive network is at least partially porous.